PubReading

PubReading [327] - DNA storage in thermoresponsive microcapsules for repeated random multiplexed data access - B. Bögels, T. de Greef et al.

Wed, 17 May 2023 11:00:40 GMT

DNA has emerged as an attractive medium for archival data storage due to its durability and high information density. Scalable parallel random access to information is a desirable property of any storage system. For DNA-based storage systems, however, this still needs to be robustly established. Here we report on a thermoconfined polymerase chain reaction, which enables multiplexed, repeated random access to compartmentalized DNA files. The strategy is based on localizing biotin-functionalized oligonucleotides inside thermoresponsive, semipermeable microcapsules. At low temperatures, microcapsules are permeable to enzymes, primers and amplified products, whereas at high temperatures, membrane collapse prevents molecular crosstalk during amplification. Our data show that the platform outperforms non-compartmentalized DNA storage compared with repeated random access and reduces amplification bias tenfold during multiplex polymerase chain reaction. Using fluorescent sorting, we also demonstrate sample pooling and data retrieval by microcapsule barcoding. Therefore, the thermoresponsive microcapsule technology offers a scalable, sequence-agnostic approach for repeated random access to archival DNA files.https://doi.org/10.1038/s41565-023-01377-4 - 2023

PubReading [346] - Transparency Is the Key to Quality - A. Fosang & R. Colbran

Mon, 03 Jul 2023 11:00:12 GMT

A workshop held last June by the National Institutes of Health (NIH) Director’s Office, Nature Publishing Group, and Science focused on the role that journals play in supporting scientific research that is reproducible, robust, and transparent. The “Principles and Guidelines for Reporting Preclinical Research” that emerged from the workshop have since been endorsed by nearly 80 societies, journals, and associations.VOL.290,NO.50,pp.29692–29694 - 2015

PubReading [343] - The changing career trajectories of new parents in STEM - E. Cech & M. Blair-Loy

Mon, 26 Jun 2023 12:00:11 GMT

The gender imbalance in science, technology, engineering, and math (STEM) fields has remained constant for decades and increases the farther up the STEM career pipeline one looks. Why does the underrepresentation of women endure? This study investigated the role of parenthood as a mechanism of gender-differentiated attrition from STEM employment. Using a nationally representative 8-year longitudinal sample of US STEM professionals, we examined the career trajectories of new parents after the birth or adoption of their first child. We found substantial attrition of new mothers: 43% of women leave full-time STEM employment after their first child. New mothers are more likely than new fathers to leave STEM, to switch to part-time work, and to exit the labor force. These gender differences hold irrespective of variation by discipline, race, and other demographic factors. However, parenthood is not just a “mother’s problem”; 23% of new fathers also leave STEM after their first child. Suggesting the difficulty of combining STEM work with caregiving responsibilities generally, new parents are more likely to leave full-time STEM jobs than otherwise similar childless peers and even new parents who remain employed full time are more likely than their childless peers to exit STEM for work elsewhere. These results have implications for policymakers and STEM workforce scholars; whereas parenthood is an important mechanism of women’s attrition, both women and men leave at surprisingly high rates after having children. Given that most people become parents during their working lives, STEM fields must do more to retain professionals with children.doi/10.1073/pnas.1810862116 - 2019

PubReading [340] - The application of human tissue for drug discovery and development - M. Clotworthy

Mon, 19 Jun 2023 12:00:12 GMT

Human tissues are invaluable resources for pharmaceutical research. They provide information about disease pathophysiology - and equally importantly, healthy function; confirmation (or refutation) of potential drug targets; validation (or otherwise) of other models employed; and functional models for assessing drugs’ effects, whether beneficial or undesirable, in the most appropriate environment that can be replicated outside the human body. While human tissues have long been prized by pathologists in furthering our under- standing of disease processes, there is a growing appreciation of their value at the late pre-clinical stage of drug discovery. Human tissues’ potential to contribute to earlier phases of the process, before significant resources have been expended, is also now gaining recognition. Mounting concern over high rates of clinical stage drug failures mandates exploration of avenues for improving efficiency. Human tissue-based assays could play a key role in improving the translation process, as well as in moving towards stratified or personalised medicines. This editorial highlights some of the potential benefits of introducing human biosamples at each stage of the research process as a drug moves from concept to clinic. Some of the challenges with respect to obtaining tissues, minimising variability and gaining acceptance are also discussed.https://doi.org/10.1517/17460441.2012.689282 - 2012

PubReading [334] - Open science challenges, benefits and tips in early career and beyond - C. Allen & D. Mehler

Mon, 05 Jun 2023 12:00:59 GMT

The movement towards open science is a consequence of seemingly pervasive failures to replicate previous research. This transition comes with great benefits but also significant challenges that are likely to affect those who carry out the research, usually early career researchers (ECRs). Here, we describe key benefits, including reputational gains, increased chances of publication, and a broader increase in the reliability of research. The increased chances of publication are supported by exploratory analyses indicating null findings are substantially more likely to be published via open registered reports in comparison to more conventional methods. These benefits are balanced by challenges that we have encountered and that involve increased costs in terms of flexibility, time, and issues with the current incentive structure, all of which seem to affect ECRs acutely. Although there are major obstacles to the early adoption of open science, overall open science practices should benefit both the ECR and improve the quality of research. We review 3 benefits and 3 challenges and provide suggestions from the perspective of ECRs for moving towards open science practices, which we believe scientists and institutions at all levels would do well to consider.https:// doi.org/10.1371/journal.pbio.3000246 - 2019

PubReading [331] - Addressing Isolation in the Scientific Community - D. Tomasello

Mon, 29 May 2023 11:00:43 GMT

In STEM, and particularly in science, many early career researchers find themselves isolated and lacking guidance. There is an enormous need to connect early career scientists with experienced professionals outside their immediate work environment. A new initiative aims to create a supportive community to foster communication between scientists through all stages of their career.https://doi.org/10.1016/j.molmed.2019.08.007 - 2019

PubReading [328] - Reflections on scientific collaboration between basic researchers and clinicians - J. Muia & C. Casari

Mon, 22 May 2023 11:00:33 GMT

Early career researchers face uncertainties with respect to their job prospects due to dwindling job markets, decreased availability of funding and undefined career paths. As basic researchers and clinicians tend to have different approaches to scientific problems, there are many advantages from successful collaborations between them. Here, we discuss how collaborations between basic and clinical scientists should be promoted early in their careers. To achieve this, researchers, both basic and clinical, must be proactive during their training and early stages of their careers. Mentors can further augment collaborative links in many ways. We suggest that universities and institutions might reassess their involvement in promoting collaborations between basic and clinical researchers. We hope that this paper will serve as a reminder of the importance of such collaborations, and provide the opportunity for all members of the scientific community to reflect on and ame- liorate their own contributions.DOI: 10.1111/jth.13447 - 2016

PubReading [325] - The Costs of Reproducibility - R. Poldrack

Mon, 15 May 2023 11:01:25 GMT

Improving the reproducibility of neuroscience research is of great concern, especially to early-career researchers (ECRs). Here I outline the potential costs for ECRs in adopting practices to improve reproducibility. I highlight the ways in which ECRs can achieve their career goals while doing better science and the need for established researchers to support them in these efforts.https://doi.org/10.1016/j.neuron.2018.11.030 - 2019

PubReading [323] - Factors that influence PhD candidates’ success: the importance of PhD project characteristics - E. van Rooij, M. Fokkens-Bruinsma & E. Jansen

Mon, 08 May 2023 13:28:12 GMT

High dropout rates, delay, and dissatisfaction among PhD students are common problems in doctoral education. Research shows that many different factors are associated with doctoral success, but these factors have not often been studied simultaneously. Moreover, characteristics of the PhD project are mostly neglected. In this study, we investigate which supervision, psychosocial, and project characteristics are related to satisfaction, progress, and quit intentions in a sample of 839 PhD candidates at a university in the Netherlands. Results of regression analyses show that experienced workload was negatively related to satisfaction and progress and positively to quit intentions. The quality of the supervisor-PhD candidate relationship, the PhD candidate’s sense of belonging, the amount of freedom in the project, and working on a project closely related to the supervisor’s research were positively related to satisfaction and negatively to quit intentions. The high workload of PhD candidates should be a major point of attention for universities who wish to increase their rates of PhD completion and PhD candidates’ satisfaction. In addition, the ‘match’ between PhD candidate and supervisor is crucial, both personally – a good relationship – and academically, i.e. that the PhD candidate works on a topic closely related to the supervisor’s research.https://doi.org/10.1080/0158037X.2019.1652158 -2021

PubReading [320] - Deep Neural Networks as Scientific Models - R. Cichy and D. Kaiser

Mon, 01 May 2023 15:19:03 GMT

Artificial deep neural networks (DNNs) initially inspired by the brain enable computers to solve cognitive tasks at which humans excel. In the absence of explanations for such cognitive phenomena, in turn cognitive scientists have started using DNNs as models to investigate biological cognition and its neural basis, creating heated debate. Here, we reflect on the case from the perspective of philosophy of science. After putting DNNs as scientific models into context, we discuss how DNNs can fruitfully contribute to cognitive science. We claim that beyond their power to provide predictions and explanations of cognitive phenomena, DNNs have the potential to contribute to an often overlooked but ubiquitous and fundamental use of scientific models: exploration.https://doi.org/10.1016/j.tics.2019.01.009 - 2019

PubReading [315] - Technology transfer programme for influenza vaccines – Lessons from the past to inform the future - C. Chadwick, E. Sparrow et al.

Mon, 27 Mar 2023 11:38:32 GMT

In 2006, to address the global inequitable access to influenza vaccines in the event of an influenza pandemic, WHO, with support of donors and partners, embarked on an ambitious project, the Technology Transfer Initiative (TTI), to facilitate influenza vaccine production capacity-building in low- and middle-income countries (LMICs). This commentary briefly summarizes the high-level lessons learned, key challenges encountered, and critical components needed for success.https://doi.org/10.1016/j.vaccine.2022.06.057 - 2022

PubReading [310] - How to Write the Methods Section of a Research Paper - R. Kallet

Mon, 20 Mar 2023 12:00:53 GMT

The methods section of a research paper provides the information by which a study’s validity is judged. Therefore, it requires a clear and precise description of how an experiment was done, and the rationale for why specific experimental procedures were chosen. The methods section should describe what was done to answer the research question, describe how it was done, justify the experimental design, and explain how the results were analyzed. Scientific writing is direct and orderly. Therefore, the methods section structure should: describe the materials used in the study, explain how the materials were prepared for the study, describe the research protocol, explain how measurements were made and what calculations were performed, and state which statistical tests were done to analyze the data. Once all elements of the methods section are written, subsequent drafts should focus on how to present those elements as clearly and logically as possibly. The description of preparations, measurements, and the protocol should be organized chronologically. For clarity, when a large amount of detail must be presented, information should be presented in sub-sections according to topic. Material in each section should be organized by topic from most to least important.49 (10) 1229-1232; - 2004

PubReading [305] - The sciences of science communication - B. Fischhoff

Mon, 13 Mar 2023 12:00:14 GMT

The May 2012 Sackler Colloquium on “The Science of Science Communication” brought together scientists with research to communicate and scientists whose research could facilitate that communication. The latter include decision scientists who can identify the scientific results that an audience needs to know, from among all of the scientific results that it would be nice to know; behavioral scientists who can design ways to convey those results and then evaluate the success of those attempts; and social scientists who can create the channels needed for trustworthy communications. This overview offers an introduction to these communication sciences and their roles in science-based communication programs.

PubReading [300] - Randomisation in clinical trials - E. Beller, V. Gebski & A. Keech

Mon, 06 Mar 2023 13:00:13 GMT

Randomisation is the process of assigning clinical trial participants to treatment groups. Randomisation gives each participant a known (usually equal) chance of being assigned to any of the groups. Successful randomisation requires that group assignment cannot be predicted in advance.

DOI: 10.5694/j.1326-5377.2002.tb04955.x - 2002

PubReading [295] - Decentralized Clinical Trials The Future of Medical Product Development? - G. Van Norman,

Mon, 27 Feb 2023 13:00:18 GMT

The Covid-19 pandemic disrupted many clinical trials that were potentially bringing new therapeutics to market—an additional untallied cost of the pandemic in lives and quality of life owing to delays in releasing potentially beneficial therapeutics to patients in need. A separate side effect of the pandemic has been swift adoption of virtual interactions between physicians and patients to provide continuity of care while maintaining social distancing. This comes at a time of rapid advancement of technology permitting those interactions, such as enhanced internet connectivity, electronic health records, real-time video conferencing, smartphone health applications, and remotely connectable health monitoring devices that are becoming both more accurate, practical, and affordable. Interest in decentralized clinical trials (DCTs) that use “virtual elements” like these has grown in parallel with acceptance of “virtual medicine,” accelerating shifts in clinical trial design that many feel are long overdue.

https://doi.org/10.1016/j.jacbts.2021.01.011 - 2021

PubReading [290] - Placebo Controls- Now??? - A. Caplan

Mon, 20 Feb 2023 13:00:17 GMT

In this era of a pandemic, why do we need the placebo controls for regulatory agency or Food and Drug Administration (FDA)-approved clinical trials aimed at COVID-19 patients? The answer is always that we have to establish a ruler with a baseline onto which efficacy measurements can be statistically judged.

https://doi.org/10.1007/s00005-021-00612-x - 2021

PubReading [285] - Promoting Scientist–Advocate Collaborations in Cancer Research- Why and How - J. Salamone, A. Shajahan-Haq et al.

Mon, 13 Feb 2023 11:00:56 GMT

Advocates bring unique and important viewpoints to the cancer research process, ensuring that scientific and medical advances are patient-centered and relevant. In this article, we discuss the benefits of engaging advocates in cancer research and underscore ways in which both the scientific and patient communities can facilitate this mutually beneficial collaboration. We discuss how to establish and nurture successful scientist-advocate relationships throughout the research process. We review opportunities that are available to advocates who want to obtain training in the evaluation of cancer research. We also suggest practical solutions that can strengthen communication between scientists and advocates, such as introducing scientist-advocate interactions at the trainee level. Finally, we highlight the essential role social media can play in disseminating patient-supported cancer research findings to the patient community and in raising awareness of the importance of promoting cancer research. Our perspective offers a model that Georgetown Breast Cancer Advocates have found effective and which could be one option for those interested in developing productive, successful, and sustainable collaborations between advocates and scientists in cancer research.

DOI: 10.1158/0008-5472.CAN-18-1600 - 2018

PubReading [280] - Ethics assessment in research proposals adopting CRISPR technology - F. Hirsch, R. Iphofen, Z. Koporc

Mon, 06 Feb 2023 12:00:19 GMT

The rapid and exponential growth of genome editing has posed many challenges for bioethics. This article briefly explains the nature of the technique and the particularly rapid development of Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) technology. The international and, specifically, European-level systems for assessing the ethical issues consequent on these developments are outlined and discussed. The challenges posed by cases in China are summarized to raise concerns about how a more shared, universally consistent appraisal of bioethical issues can be promoted.

https://doi.org/10.11613/BM.2019.020202 - 2019

PubReading [275] - The biospheric emergency calls for scientists to change tactics - F. Racimo, J. Halder et al.

Mon, 30 Jan 2023 12:01:26 GMT

Our current economic and political structures have an increasingly devastating impact on the Earth's climate and ecosystems: we are facing a biospheric emergency, with catastrophic consequences for both humans and the natural world on which we depend. Life scientists - including biologists, medical scientists, psychologists and public health experts - have had a crucial role in documenting the impacts of this emergency, but they have failed to drive governments to take action in order to prevent the situation from getting worse. Here we, as members of the movement Scientist Rebellion, call on life scientists to re-embrace advocacy and activism - which were once hallmarks of academia - in order to highlight the urgency and necessity of systemic change across our societies. We particularly emphasise the need for scientists to engage in nonviolent civil resistance, a form of public engagement which has proven to be highly effective in social struggles throughout history.

doi: 10.7554/eLife.83292 - 2022

PubReading [270] - Why do researchers decide to publish in questionable journals? A review of the literature - T. Frandsen

Mon, 23 Jan 2023 13:00:23 GMT

Peer review is a cornerstone of scientific publication, and consequently, predatory journals are feared to be a threat to the credibility of science as they perform no or low-quality peer review. The question of why researchers decide to publish in a questionable journal remains relatively unexplored. This paper provides an overview of the existing literature on why researchers decide to publish papers in questionable journals, specifically whether or not they search for a low-barrier way to getting published while being aware that the chosen journal probably does not adhere to acceptable academic standards. The choice of a publication outlet can be seen as a submission tree that consists of various incentives, and explaining why authors publish in deceptive journals may thus consist of a combination of awareness and motivational factors. Awareness and motivation of diligent authors is very different from that of unethical authors. Unethical authors may use a lack of awareness to excuse their actions, but they may actively search for a low-barrier way to getting published. As there are different types of authors who publish in deceptive journals, we need different approaches to solve the problem.

doi: 10.1002/leap.1214 - 2018

PubReading [265] - The art of writing literature review- What do we know and what do we need T to know? - J. Paula & A. Criado

Mon, 16 Jan 2023 13:00:12 GMT

A literature review article provides a comprehensive overview of literature related to a theme/theory/method and synthesizes prior studies to strengthen the foundation of knowledge. In the growing International Business (IB) research field, systematic literature reviews have great value, yet there are not many reviews published describing how researchers can design and develop classic review articles. In explaining the purpose, methodology, and structure of a systematic review, we provide guidelines for developing most insightful and useful review articles. By outlining steps and thumb rules to keep in mind, we present an overview of different types of review articles and explain how future researchers could potentially find them useful. In addition, we introduce nine articles finally selected for this special issue of systematic literature review-Looking back to look forward International Business research in the days to come.

https://doi.org/10.1016/j.ibusrev.2020.101717 - 2020

PubReading [260] - Towards a framework for embedding doctorateness in research proposals - E. Sanganyado, W. Nunu and S. Sanganyado

Mon, 09 Jan 2023 12:00:34 GMT

Doctorate graduates are expected to contribute original knowledge and possess advanced skills essential for addressing complex problems. Embedding doctorateness in doctorate programmes could help ensure that the productivity of doctoral research is explicitly demonstrated. Doctorateness represents independent scholarship, the transition from knowledge consumption to knowledge creation, original scholarly contribution, and research integrity. A research proposal is the backbone of a doctoral journey, as it acts as an academic development plan for supervisors and students. Hence, understanding the role of supervisors in enabling doctorateness in preparing a research proposal is essential for improving student satisfaction and competencies in their academic development. Here, we develop a framework for facilitating doctorateness in preparing a research proposal and provide recommen- dations on supervisor functions.

https://doi.org/10.1080/14703297.2022.2124186 - 2022

PubReading [255] - Doing better- eleven ways to improve the integration of sex and gender in health research proposals - R. Mason

Mon, 02 Jan 2023 12:00:32 GMT

Health researchers are often challenged on how to integrate a sex and gender lens into their work. Reviews completed across a range of health research studies show there are several commonly overlooked opportunities to do better in this regard. Nine ways to improve the integration of a sex and gender lens in health research proposals have been identified.

https://doi.org/10.1186/s41073-020-00102-2 - 2020

PubReading [240] - Is my paper relevant for an international audience? - F. Fernandez-llimos

Mon, 12 Dec 2022 12:00:57 GMT

This is the first question one should consider before submitting a paper to an international journal. The answer is simple: If researchers or practitioners from another country can learn something from your paper that can influence a practice or a research they are involved in, then your paper is relevant for an international audience. There are many elements that can influence in this cross-border transferability. One could think that having a big “n”, or performing complex statistical calculations, or using complicated study designs makes the paper more attractive to colleagues from other countries. These elements can help, but they are not sufficient. On the other hand, one could think that a study performed in a small hospital in a given country will never be of interest for these foreign colleagues. That is not necessarily correct. Let’s burst some myths.

https://doi.org/10.18549/PharmPract.2020.2.1924 - 2020

PubReading [235] - How to...write a good research question - K. Mattick, J. Johnston and A. de la Croix

Mon, 05 Dec 2022 12:00:13 GMT

This paper, on writing research questions, is the first in a series that aims to support novice researchers within clinical education, particularly those undertaking their first qualitative study. Put simply, a research question is a question that a research project sets out to answer. Most research questions will lead to a project that aims to generate new insights, but the target audience and the methodology will vary widely. The term ‘evaluation question’ is used less commonly, but the same principles apply. The key difference is that evaluation questions are typically more focused on the immediate context: for example, the effectiveness of an educational intervention in a particular setting. Whether your ambition is for research or evaluation, we hope that you will find this paper helpful for designing your own educational projects.

DOI: doi: 10.1111/tct.12776 - 2018

PubReading [230] - Does a Graphical Abstract Bring More Visibility to Your Paper? - E. Pferschy-Wenzig, A. Atanasov et al.

Mon, 28 Nov 2022 12:00:17 GMT

A graphical abstract (GA) represents a piece of artwork that is intended to summarize the main findings of an article for readers at a single glance. Many publishers currently encourage authors to supplement their articles with GAs, in the hope that such a convenient visual summary will facilitate readers with a clearer outline of papers that are of interest and will result in improved overall visibility of the respective publication. To test this assumption, we statistically compared publications with or without GA published in Molecules between March 2014 and March 2015 with regard to several output parameters reflecting visibility. Contrary to our expectations, manuscripts published without GA performed significantly better in terms of PDF downloads, abstract views, and total citations than manuscripts with GA. To the best of our knowledge, this is the first empirical study on the effectiveness of GA for attracting attention to scientific publications.

doi:10.3390/molecules21091247 - 2016

PubReading [225] - A brief guide to the science and art of writing manuscripts in biomedicine - Di. Forero, S. Lopez-Leon, G. Perry

Mon, 21 Nov 2022 12:00:18 GMT

Publishing articles in international scientific journals is the primary method for the communication of validated research findings and ideas. Journal articles are commonly used as a major input for evaluations of researchers and institutions. Few articles have been published previously about the different aspects needed for writing high-quality articles. In this manuscript, we provide an updated and brief guide for the multiple dimensions needed for writing manuscripts in the health and biological sciences, from current, international and interdisciplinary perspectives and from our expertise as authors, peer reviewers and editors. We provide key suggestions for writing major sections of the manuscript (e.g. title, abstract, introduction, methods, results and discussion), for submitting the manuscript and bring an overview of the peer review process and of the post-publication impact of the articles.

https://doi.org/10.1186/s12967-020-02596-2 - 2022

PubReading [190] - Contextualizing the Impostor “Syndrome” - S. Feenstra, J. Jordan et al.

Mon, 03 Oct 2022 20:00:18 GMT

The impostor “syndrome” refers to the notion that some individuals feel as if they ended up in esteemed roles and positions not because of their competencies, but because of some oversight or stroke of luck. Such individuals therefore feel like frauds or “impostors.” Despite the fact that impostor feelings are often linked to marginalized groups in society, to date, research predominantly approaches this phenomenon as an issue of the individual: pointing toward individuals for the roots and solutions of the “syndrome.” Drawing from a rich body of social and organizational psychology research, in this perspectives piece, we propose a shift in how scholars conceptualize and empirically examine this phenomenon. Instead of framing the insecurities of individuals belonging to marginalized groups solely as a problem that arises within these individuals, we argue that it is critical for future research to consider the important role of the environment in eliciting their impostor feelings as well. By doing so, we can address the contextual roots of individuals’ impostor feelings, and offer more structural and effective solutions.

doi: 10.3389/fpsyg.2020.575024 - 2020

PubReading [47] - Renal artery aneurysms - D. M. Coleman and J. C. Stanley

Wed, 27 Oct 2021 20:00:22 GMT

Renal artery aneurysms are rare in the general population, although the true incidence and natural history remain elusive. Controversy over criteria for repair persists across decades. Indications for repair presently include aneurysm size >2 cm, female gender within childbearing age, symptoms like pain and hematuria, medically refractory hypertension including that associated with functionally important renal artery stenosis, thromboembolism, dissection, and rupture. Conventional surgical reconstruction options are variable and continue to offer technically sound and durable results. Endovascular therapies with novel devices also offer technical success with few major adverse events, and are increasingly employed as indications for intervention broaden. This review summarizes the accumulated evidence on true renal artery aneurysms with a particular focus on contemporary treatment criteria, natural history, options for repair and outcomes following such. -doi.org/10.1016/j.jvs.2015.05.034 - 2015

PubReading [20] - Anatomy, Back, Cauda Equina - E. Berg, J. Ashurst.

Mon, 06 Sep 2021 22:02:15 GMT

In 1595, French anatomist Andre du Laurens first described the structure of a rope-like tail of fibers at the caudal end of the spinal cord. This bundle of numerous axons was termed the cauda equina, from the Latin translation meaning “horse’s tail,” and it contains nerves which innervate both sensory and motor targets within lumbar, sacral, and coccygeal spinal cord levels. Epidemiologic assessments regard lesions to the cauda equina as uncommon, with a prevalence of 1 to 3 per 100,000 subjects. Typically caused by a herniated intervertebral disc at the L5-S1 levels, such lesions affect females as often as males and manifest as a number of urogenital and neuromuscular symptoms in the namesake “cauda equina syndrome." - StatPearls Publishing -2021

PubReading [19] - Hospitalization Length after Myocardial Infarction: Risk-Assessment-Based Time of Hospital Discharge vs. Real Life Practice - Michał Wegiel and Tomasz Rakowski

Thu, 02 Sep 2021 22:13:27 GMT

According to guidelines, it is safe for low-risk patients with myocardial infarction (MI) to be discharged within 72 h of hospitalization. However, results coming from registries show that the hospital stay is often much longer in a real-life situation. Data on the length of the hospital stay (LOS) of MI patients in Polish centers are lacking. We enrolled 212 consecutive patients with acute MI. Low-risk patients were defined according to PAMI II criteria: age 45%, no persistent ventricular arrhythmia, and no multi-vessel disease (MVD). The median of the hospitalization length was eight days (Q1: 6; Q3: 9). In low-risk patients (25%), the median of LOS was six days (Q1: 5; Q3: 7) (p < 0.001). In a logistic regression analysis patients age, LVEF, ST-segment-elevation MI and the presence of MVD were independent predictors of longer hospitals stay (≥8 days). During follow up, there were no significant differences in the rates of clinical events between patients with shorter (

PubReading [14] - Acute abdominal pain in patients with Crohn’s disease: What urgent imaging tests should be done? - P. García, A. López-Jurado, A. Bártulos

Fri, 20 Aug 2021 23:42:04 GMT

Crohn’s disease is an autoimmune disease that predominantly affects the gastrointestinal tract. Crohn’s disease is diagnosed at a young age and runs a chronic course with acute flare-ups. When patients with Crohn’s disease present with flare-ups at the emergency department, they are usually managed in a way similar to patients with acute abdomen; there is no consensus about the most appropriate imaging work-up for patients with flare-ups of Crohn’s disease. Thus, we decided to review the literature about the imaging tests indicated (whether related to their diagnostic performance or to lower exposure to ionizing radiation) for acute flare-ups in patients with Crohn’s disease. - DOI: 10.1016/j.rx.2018.12.003 - 2019

PubReading [11] - Outcomes of critically ill patients with acute kidney injury in COVID-19 infection: an observational study - Rodrigo Bezerra et al.

Tue, 17 Aug 2021 20:16:10 GMT

Background: Early reports indicate that AKI is common during COVID-19 infection. Different mortality rates of AKI due to SARS-CoV-2 have been reported, based on the degree of organic dysfunction and varying from public to private hospitals. However, there is a lack of data about AKI among critically ill patients with COVID-19.

Methods: We conducted a multicenter cohort study of 424 critically ill adults with severe acute respiratory syndrome (SARS) and AKI, both associated with SARS-CoV-2, admitted to six public ICUs in Brazil. We used multivariable logistic regression to identify risk factors for AKI severity and in-hospital mortality.

Results: The average age was 66.42±13.79years, 90.3% were on mechanical ventilation (MV), 76.6% were at KDIGO stage 3, and 79% underwent hemodialysis. The overall mortality was 90.1%. We found a higher frequency of dialysis (82.7% versus 45.2%), MV (95% versus 47.6%), vasopressors (81.2% versus 35.7%) (p

PubReading [10] - Recommendations on pre-hospital and early hospital management of acute heart failure - short version - Alexandre Mebazaa et al.

Thu, 12 Aug 2021 23:16:31 GMT

Despite several critical steps forward in the management of chronic heart failure (CHF), the area of acute heart failure (AHF) has remained relatively stagnant. As stated in the updated ESC HF guidelines, clinicians responsible for managing patients with AHF must frequently make treatment decisions without adequate evidence, usually on the basis of expert opinion consensus.2 Specifically, the treatment of acute HF remains largely opinion-based with little good evidence to guide therapy. Acute heart failure is a syndrome in which emergency physicians, cardiologists, intensivists, nurses, and other healthcare providers have to cooperate to provide ‘rapid’ benefit to the patients. We hereby would like to underscore the wider experience grown in different settings of the area of intensive care on acute heart failure, actually larger and more composite than that got in specialized Care Units. The distillate of such different experiences is discussed and integrated in the present document. Hence, the authors of this consensus paper believe a common working definition of AHF covering all dimensions and modes of presentations has to be made, with the understanding that most AHF presentations are either acute decompensations of chronic underlying HF or the abrupt onset of dyspnoea associated with significantly elevated blood pressure. Secondly, recent data show that, much like acute coronary syndrome, AHF might have a ‘time to therapy’ concept. Accordingly, ‘pre-hospital’ management is considered a critical component of care. Thirdly, most patients with AHF have normal or high blood pressure at presentation and are admitted with symptoms and/or signs of congestion. This is in contradiction to the presentation where low cardiac output leads to symptomatic hypotension and signs/symptoms of hypoperfusion, a circumstance that is relatively rare, present in coronary care unit/ intensive care unit (CCU/ICU) but associated with a particularly poor outcome. Hence, it is important to note that appropriate therapy requires appropriate identification of the specific AHF pheno-type.3 The aim of the current paper is not to replace guidelines, but, to provide contemporary perspective for early hospital management within the context of the most recent data and to provide guidance, based on expert opinions, to practicing physicians and other healthcare professionals (Figure 1). We believe that the experience accrued in the different settings from the emergency department through to the ICU/CCU is collectivel valuable in determining how best to manage the patients with AHF. Herein, a shortened version mainly including group recommendations is provided. Full version of the consensus paper is provided as Supplementary material online. - Alexandre Mebazaa - http://onlinelibrary.wiley.com/doi/10.1002/ejhf.289/full - 2015

PubReading [341] - Carbon Nanomaterials (CNMs) in Cancer Therapy- A Database of CNM-Based Nanocarrier Systems - H. Mohan, A. Fagan & S. Giordani

Wed, 21 Jun 2023 11:00:12 GMT

Carbon nanomaterials (CNMs) are an incredibly versatile class of materials that can be used as scaffolds to construct anticancer nanocarrier systems. The ease of chemical functionalisation, biocompatibility, and intrinsic therapeutic capabilities of many of these nanoparticles can be leveraged to design effective anticancer systems. This article is the first comprehensive review of CNM-based nanocarrier systems that incorporate approved chemotherapy drugs, and many different types of CNMs and chemotherapy agents are discussed. Almost 200 examples of these nanocarrier systems have been analysed and compiled into a database. The entries are organised by anticancer drug type, and the composition, drug loading/release metrics, and experimental results from these systems have been compiled. Our analysis reveals graphene, and particularly graphene oxide (GO), as the most frequently employed CNM, with carbon nanotubes and carbon dots following in popularity. Moreover, the database encompasses various chemotherapeutic agents, with antimicrotubule agents being the most common payload due to their compatibility with CNM surfaces. The benefits of the identified systems are discussed, and the factors affecting their efficacy are detailed.doi.org/10.3390/pharmaceutics15051545 - 2023

PubReading [339] - Pleiotropic Effects of Icariside II on the Cardiovascular System- Novel Applications of Ethnopharmacology in Targeting Vascular Remodeling - C. Shan, R. Murphy et al.

Fri, 16 Jun 2023 12:00:13 GMT

Albert Einstein once eloquently stated “We still do not know one thousandth of one percent of what nature has revealed to us.” Mother Nature has proven time and again to be the best engineer, architect, scientist, and doctor. This fact has not been lost on mankind, and since time immemorial, we have looked to nature for answers to human conditions and diseases. Indeed, our first evidence dates back to preliterary history as long as 50,000 years ago, in archaeological discoveries from a Middle Eastern grave site at Shanidar, Iraq, of a Neanderthal man which contained plant specimens, of which are still used in local traditional medicine. Since then, the use and application of herbal medicine has been recorded in every society from Traditional Chinese, Ayurvedic, Yunani, and other cultures in the developing world.DOI: 10.1097/FJC.0000000000001278 - 2022

PubReading [338] - A Systems Biology Approach to Investigate Kinase Signal Transduction Networks That Are Involved in Triple Negative Breast Cancer Resistance to Cisplatin - N. Mukherjee, A. Eustace et al.

Wed, 14 Jun 2023 12:00:14 GMT

Triple negative breast cancer (TNBC) remains a therapeutic challenge due to the lack of targetable genetic alterations and the frequent development of resistance to the standard cisplatin-based chemotherapies. Here, we have taken a systems biology approach to investigate kinase signal transduction networks that are involved in TNBC resistance to cisplatin. Treating a panel of cisplatin- sensitive and cisplatin-resistant TNBC cell lines with a panel of kinase inhibitors allowed us to reconstruct two kinase signalling networks that characterise sensitive and resistant cells. The analysis of these networks suggested that the activation of the PI3K/AKT signalling pathway is critical for cisplatin resistance. Experimental validation of the computational model predictions confirmed that TNBC cell lines with activated PI3K/AKT signalling are sensitive to combinations of cisplatin and PI3K/AKT pathway inhibitors. Thus, our results reveal a new therapeutic approach that is based on identifying targeted therapies that synergise with conventional chemotherapies.doi.org/10.3390/jpm12081277 - 2022

PubReading [336] - Foldaxanes- Rotaxane-like Architectures from Foldamers - V. Koegler, Y. Ferrand et al.

Fri, 09 Jun 2023 12:00:40 GMT

Mechanically interlocked molecules such as rotaxanes and catenanes contain free-moving components that cannot dissociate and have enabled the investigation and control of various translational and rotational molecular motions. The architecture of pseudo-rotaxanes and of some kinetically labile rotaxanes is comparable to that of rotaxanes but their components are reversibly associated and not irreversibly interlocked. In other words, pseudo-rotaxanes may fall apart. This Account focuses on a peculiar family of rotaxane-like architectures termed foldaxanes. Foldaxanes consist of a helically folded oligomer wound around a rod-like dumbbell-shaped guest. Winding of the helix around the rod thus entails an unwinding−rewinding process that creates a kinetic barrier. It follows that foldaxanes, albeit reversibly assembled, have significant lifetimes and may not fall apart while defined molecular motions are triggered. Foldaxanes based on helically folded aromatic oligoamide hosts and oligo(alkyl carbamate) guests can be designed rationally through the inclusion of complementary binding motifs on the rod and at the inner rim of the helix so that helix length and rod length match. Single helical foldaxanes (bimolecular species) and double helical foldaxanes (trimolecular species) have thus been produced as well as poly[n]foldaxanes, in which several helices bind to long rods with multiple binding stations. When the binding stations differ and are organized in a certain sequence, a complementary sequence of different stacked helices, each matching with their binding station, can be assembled, thus reproducing in an artificial system a sort of translation process.https://doi.org/10.1021/acs.accounts.2c00050 - 2022

PubReading [335] - Delta chirality ruthenium ‘light-switch’ complexes can bind in the minor groove of DNA with five different binding modes - J. Hall, C. Cardin et al.

Wed, 07 Jun 2023 12:00:42 GMT

[Ru(phen)2 (dppz)]2+ has been studied since the 1990s due to its ‘light-switch’ properties. It can be used as a luminescent DNA probe, with emission switched on through DNA binding. The luminescence observed is dependent on the solvent accessibility of the pyrazine nitrogen atoms, and therefore is sensitive to changes in both binding site of the cation and chromophore orientation. The compound is also chiral, and there are distinct differences between the enantiomers in terms of the emission behaviour when bound to a variety of DNA sequences. Whilst a number of binary DNA-complex X-ray crystal structures are available, most include the lambda enantiomer and there is very little structural information about binding of the delta enantiomer. Here, we present the first X-ray crystal structure of a delta enantiomer bound to well-matched DNA, in the absence of the other, lamda enantiomer. We show how the binding site observed here can be related to a more general pattern of motifs in the crystallographic literature and propose that the delta enantiomer can bind with five different binding modes, offering a new hypothesis for the interpretation of solution data.doi: 10.1093/nar/gkw753 - 2016

PubReading [333] - Enzymatic assembly of DNA molecules up to several hundred kilobases - D. Gibson, H. Smith et al.

Wed, 31 May 2023 11:40:13 GMT

We describe an isothermal, single-reaction method for assembling multiple overlapping DNA molecules by the concerted action of a 5' exonuclease, a DNA polymerase and a DNA ligase. First we recessed DNA fragments, yielding single-stranded DNA overhangs that specifically annealed, and then covalently joined them. This assembly method can be used to seamlessly construct synthetic and natural genes, genetic pathways and entire genomes, and could be a useful molecular engineering tool.DOI:10.1038/NMETH.1318 - 2009

PubReading [332] - Do natural DNA triple-helical structures occur and function in vivo? -

Tue, 30 May 2023 11:00:16 GMT

Formation of natural intramolecular triple-helical structures of DNA is still an intriguing research topic in view of the possible involvement of these structures in biological processes. The biochemical and biophysical properties of DNA triplex structures have been extensively studied, and experimental data show that H-DNA is likely to form in vivo and may regulate the expression of various genes. However, direct and unambiguous evidence of the possible biological roles of these structures is yet elusive. This review focuses on the basic facts that are in favor of, or against, the hypothesis of the presence and function of natural DNA triple-helical structures in vivo, and outlines the different methods and probes that have been used to support these facts.DOI 10.1007/s00018-003-3046-3 - 2003

PubReading [330] - Diversity and Complexity of the Large Surface Protein Family in the Compacted Genomes of Multiple Pneumocystis Species - L. Ma, J. Kovacs et al.

Wed, 24 May 2023 12:00:58 GMT

Pneumocystis, a major opportunistic pathogen in patients with a broad range of immunodeficiencies, contains abundant surface proteins encoded by a multicopy gene family, termed the major surface glycoprotein (Msg) gene superfamily. This superfamily has been identified in all Pneumocystis species characterized to date, highlighting its important role in Pneumocystis biology. In this report, through a comprehensive and in-depth characterization of 459 msg genes from 7 Pneumocys- tis species, we demonstrate, for the first time, the phylogeny and evolution of conserved domains in Msg proteins and provide a detailed description of the classification, unique characteristics, and phylogenetic relatedness of five Msg families. We further describe, for the first time, the relative expression levels of individual msg families in two rodent Pneumocystis species, the substantial variability of the msg repertoires in P. carinii from laboratory and wild rats, and the distinct features of the expression site for the classic msg genes in Pneumocystis from 8 mammalian host species. Our analysis suggests multiple functions for this superfamily rather than just conferring antigenic variation to allow immune evasion as previously believed. This study provides a rich source of information that lays the foundation for the continued experimental exploration of the functions of the Msg superfamily in Pneumocystis biology.https://doi.org/10 .1128/mBio.02878-19 - 2020

PubReading [329] - The immune suppressive tumor microenvironment in multiple myeloma- The contribution of myeloid-derived suppressor cells - C. Giannotta, F. Autino and M. Massaia

Tue, 23 May 2023 11:00:35 GMT

Myeloid derived suppressors cells (MDSC) play major roles in regulating immune homeostasis and immune responses in many conditions, including cancer. MDSC interact with cancer cells within the tumor microenvironment (TME) with direct and indirect mechanisms: production of soluble factors and cytokines, expression of surface inhibitory molecules, metabolic rewiring and exosome release. The two-way relationship between MDSC and tumor cells results in immune evasion and cancer outgrowth. In multiple myeloma (MM), MDSC play a major role in creating protumoral TME conditions. In this minireview, we will discuss the interplay between MDSC and MM TME and the possible strategies to target MDSC.doi: 10.3389/fimmu.2022.1102471 - 2023

PubReading [326] - Drug screening for human genetic diseases using iPSC models - M. Elitt, L. Barbar & P. Tesar

Tue, 16 May 2023 11:40:19 GMT

Induced pluripotent stem cells (iPSCs) enable the generation of previously unattainable, scalable quantities of disease- relevant tissues from patients suffering from essentially any genetic disorder. This cellular material has proven instrumental for drug screening efforts on these disorders, and has facilitated the identification of novel therapeutics for patients. Here we will review the foundational technologies that have enabled iPSCs, the power and limitations of iPSC-based compound screens along with screening guidelines, and recent examples of screening efforts. Additionally we will provide a brief commentary on the future scientific roadmap using pluripotent- and 3D organoid-based, combinatorial approaches.doi: 10.1093/hmg/ddy186 - 2018

PubReading [325] - pH-dependent sedimentation of DNA in the presence of divalent, but not monovalent, metal ions - C. England, L. K. Lewis et al.

Wed, 10 May 2023 11:01:32 GMT

Precipitation of DNA is performed frequently in molecular biology laboratories for the purpose of purification and concentration of samples and also for transfer of DNA into cells. Metal ions are used to facilitate these processes, though their precise functions are not well characterized. In the current study we have investigated the precipitation of double-stranded DNA by group 1 and group 2 metal ions. Double-stranded DNAs were not sedimented efficiently by metals alone, even at high concentrations. Increasing the pH to 11 or higher caused strong DNA precipitation in the presence of the divalent group 2 metals magnesium, calcium, strontium and barium, but not group 1 metals. Group 2 sedimentation profiles were distinctly different from that of the transition metal zinc, which caused precipitation at pH 8. Analysis of DNAs recovered from precipitates formed with calcium revealed that structural integrity was retained and that sedimentation efficiency was largely size-independent above 400 bp. Several tests supported a model whereby single-stranded DNA regions formed by denaturation at high pH became bound by the divalent metal cations. Neutralization of negative surface charges reduced the repulsive forces between molecules, leading to formation of insoluble aggregates that could be further stabilized by cation bridging (ionic crosslinking).doi:10.1016/j.ab.2020.114099 - 2021

PubReading [324] - DNA sequencing with chain-terminating inhibitors - F. Sanger, S.Nickel & A. Coulson

Tue, 09 May 2023 11:01:32 GMT

A new method for determining nucleotide sequences in DNA is described. It is similar to the "plus and minus" method but makes use of the 2',3 '- dideoxy and arabinonucleoside analogues of the normal deoxynucleoside triphosphates, which act as specific chain-terminating inhibitors of DNA polymerase. The technique has been applied to the DNA of bacteriophage phiX174 and is more rapid and more accurate than either the plus or the minus method.doi: 10.1073/pnas.74.12.5463. - 1977

PubReading [322] - Millisecond-timescale, genetically targeted optical control of neural activity - E. Boyden, K. Deisseroth et al.

Wed, 03 May 2023 11:00:58 GMT

Temporally precise, noninvasive control of activity in well-defined neuronal populations is a long-sought goal of systems neuroscience. We adapted for this purpose the naturally occurring algal protein Channelrhodopsin-2, a rapidly gated light-sensitive cation channel, by using lentiviral gene delivery in combination with high-speed optical switching to photostimulate mammalian neurons. We demonstrate reliable, millisecond- timescale control of neuronal spiking, as well as control of excitatory and inhibitory synaptic transmission. This technology allows the use of light to alter neural processing at the level of single spikes and synaptic events, yielding a widely applicable tool for neuroscientists and biomedical engineers.doi:10.1038/nn1525 - 2005

PubReading [321] - Controlling the double helix - G. Felsenfeld & M. Groudine

Tue, 02 May 2023 07:39:55 GMT

Chromatin is the complex of DNA and proteins in which the genetic material is packaged inside the cells of organisms with nuclei. Chromatin structure is dynamic and exerts profound control over gene expression and other fundamental cellular processes. Changes in its structure can be inherited by the next generation, independent of the DNA sequence itself.doi:10.1038/nature01411 - 2003

PubReading [319] - The Science Behind NMN—A Stable, Reliable NAD+ Activator and Anti-Aging Molecule - Christopher Shade

Fri, 31 Mar 2023 11:00:30 GMT

In June of 2018, the World Health Organization (WHO) released the 11th edition of its International Classification of Diseases, and for the first time added aging. The classification of aging as a disease paves the way for new research into novel therapeutics to delay or reverse age-related illnesses such as cancer, cardiovascular and metabolic disease, and neurodegeneration. Nutrient sensing systems have been an intense focus of investigation, including mTOR (the mammalian target of rapamycin) for regulating protein synthesis and cell growth; AMPK (activated protein kinase) for sensing low energy states; and sirtuins, a family of seven proteins critical to DNA expression and aging, which can only function in conjunction with NAD+ (nicotinamide adenine dinucleotide), a coenzyme present in all living cellsIntegr Med (Encinitas).2020 Feb;19(1):12-14.

PubReading [318] - The essential metals for humans- a brief overview - M. Zoroddua, V. Nurchi et al.

Thu, 30 Mar 2023 11:00:35 GMT

The human body needs about 20 essential elements in order to function properly and among them, for certain, 10 are metal elements, though for every metal we do need, there is another one in our body we could do without it. Until about 1950 poor attention was given to the so-called “inorganic elements” and while researches on “organic elements” (C, N, O and H) and organic compounds were given high priority, studies on essential inorganic elements were left aside. Base on current knowledge it is ascertained today that metals such as Na, K, Mg, Ca, Fe, Mn, Co, Cu, Zn and Mo are essential elements for life and our body must have appropriate amounts of them. Here a brief overview to highlight their importance and current knowledge about their essentiality.https://doi.org/10.1016/j.jinorgbio.2019.03.013 - 2019

PubReading [317] - Nucleic acid sensing via electrochemical oligonucleotide-templated reactions - P. Gillespie, D. O’Har et al.

Wed, 29 Mar 2023 11:00:43 GMT

Short single-stranded nucleic acids as found in a variety of bodily fluids have recently emerged as minimally invasive biomarkers for a broad range of pathologies, most notably cancer. Because of their small size, low natural abundance and high sequence homology between family members they are challenging to detect using standard technologies suitable for use at the point-of-care. Herein we report the design, engineering and testing of a novel sensing strategy: electrochemically active molecular probes based on peptide nucleic acid (PNA) scaffolds for the detection of single-stranded oligonucleotides, in particular microRNAs (or miRs). As a proof-of- principle, a wide range of probes were designed and tested to detect miR-141, a known diagnostic biomarker for prostate cancer. Optimal quantitative sensing of miR-141 was achieved via the first example of an electro-chemical oligonucleotide-templated reaction (EOTR), whereby two PNA probes - functionalized with an aniline and a 1,4-catechol respectively - preferentially react with each other upon simultaneous hybridization to the same RNA target strand, serving here as a template. Quantitative, electrochemical detection of the product of this bio-orthogonal reaction showed direct correlation between adduct formation and miR-141 concentration. Coupling the specificity of OTR with the speed and sensitivity of electrochemical sensing delivers EOTRs as a promising new technique for fast, low-cost, quantitative and sequence-specific detection of short nucleic acids from liquid biopsies.https://doi.org/10.1016/j.bios.2020.112891 - 2020

PubReading [316] - A brief history of human disease genetics - M. Claussnitzer, M. McCarthy et al.

Tue, 28 Mar 2023 11:00:53 GMT

A primary goal of human genetics is to identify DNA sequence variants that influence biomedical traits, particularly those related to the onset and progression of human disease. Over the past 25 years, progress in realizing this objective has been transformed by advances in technology, foundational genomic resources and analytical tools, and by access to vast amounts of genotype and phenotype data. Genetic discoveries have substantially improved our understanding of the mechanisms responsible for many rare and common diseases and driven development of novel preventative and therapeutic strategies. Medical innovation will increasingly focus on delivering care tailored to individual patterns of genetic predisposition.https://doi.org/10.1038/s41586-019-1879-7 - 2020

PubReading [314]- MicroRNAs, damage levels, and DNA damage response control - H. Visser and A. Thomas

Fri, 24 Mar 2023 12:00:26 GMT

DNA damage–inducible miRNAs are likely to be functional in the DNA damage response. This response can elicit damage resolution and cell survival or apoptosis. The current, albeit incomplete, picture suggests that miRNAs can affect cell fate via modulation of key response proteins, but the question is, who’s in charge?https://doi.org/10.1016/j.tig.2021.06.018 - 2021

PubReading [313] - Physical activity, exercise, and chronic diseases- A brief review - E. Anderson, J. L. Durstine

Thu, 23 Mar 2023 12:00:26 GMT

Chronic diseases are the leading cause of death worldwide with increasing prevalence in all age groups, genders, and ethnicities. Most chronic disease deaths occur in middle-to low-income countries but are also a significant health problem in developed nations. Multiple chronic diseases now affect children and adolescents as well as adults. Being physically inactive is associated with increased chronic disease risk. Global societies are being negatively impacted by the increasing prevalence of chronic disease which is directly related to rising healthcare expenditures, workforce complications regarding attendance and productivity, military personnel recruitment, and academic success. However, increased physical activity (PA) and exercise are associated with reduced chronic disease risk. Most physiologic systems in the body benefit positively from PA and exercise by primary disease prevention and secondary disease prevention/treatment. The purpose of this brief review is to describe the sig- nificant global problem of chronic diseases for adults and children, and how PA and exercise can provide a non-invasive means for added prevention and treatment.https://doi.org/10.1016/j.smhs.2019.08.006 - 2019

PubReading [312] - Identification of endocrine disrupting chemicals acting on human aromatase - R. Baravalle, G. Gilardi et al.

Wed, 22 Mar 2023 12:00:44 GMT

Human aromatase is the cytochrome P450 catalysing the conversion of androgens into estrogens playing a key role in the endocrine system. Due to this role, it is likely to be a target of the so-called endocrine disrupting chemicals, a series of compounds able to interfere with the hormone system with toxic effects. If on one side the toxicity of some compounds such as bisphenol A is well known, on the other side the toxic concentrations of such compounds as well as the effect of the many other molecules that are in contact with us in everyday life still need a deep investigation. The availability of biological assays able to detect the interaction of chemicals with key molecular targets of the endocrine system represents a possible solution to identify potential endocrine disrupting chemicals.Here the so-called alkali assay previously developed in our laboratory is applied to test the effect of different compounds on the activity of human aromatase. The assay is based on the detection of the alkali product that forms upon strong alkali treatment of the NADP+ released upon enzyme turnover. Here it is applied on human aromatase and validated using anastrozole and sildenafil as known aromatase inhibitors. Out of the small library of compounds tested, resveratrol and ketoconazole resulted to inhibit aromatase activity, while bisphenol A and nicotine were found to exert an inhibitory effect at relatively high concentrations (100 μM), and other molecules such as lindane and four plasticizers did not show any significant effect. These data are confirmed by quantification of the product estrone in the same reaction mixtures through ELISA.Overall, the results show that the alkali assay is suitable to screen for molecules that interfere with aromatase activity. As a consequence it can also be applied to other molecular targets of EDCs that use NAD(P)H for catalysis in a high throughput format for the fast screening of many different compounds as endocrine disrupting chemicals.http://dx.doi.org/10.1016/j.bbapap.2017.05.013 - 2017

PubReading [311] - Chemical biology of non-canonical structures of nucleic acids for therapeutic applications - H. Tateishi-Karimata and N. Sugimoto

Tue, 21 Mar 2023 12:00:23 GMT

DNA forms not only the canonical duplex structure but also non-canonical structures. Most potential sequences that induce the formation of non-canonical structures are present in disease-related genes. Interestingly, biological reactions are inhibited or dysregulated by non-canonical structure formation in disease-related genes. To control biological reactions, methods for inducing the formation of non- canonical structures have been developed using small molecules and oligonucleotides. In this feature article, we review biological reactions such as replication, transcription, and reverse transcription controlled by non-canonical DNA structures formed by disease-related genes. Furthermore, we discuss recent studies aimed at developing methods for regulating these biological reactions using drugs targeting the DNA structure.DOI: 10.1039/c9cc09771f - 2020

PubReading [309] - What We Talk About When We Talk About “Junk DNA” - N. Fagundes, A. Zani et al.

Fri, 17 Mar 2023 12:00:47 GMT

“Junk DNA” is a popular yet controversial concept that states that organisms carry in their genomes DNA that has no positive impact on their fitness. Nonetheless, biochemical functions have been identified for an increasing fraction of DNA elements traditionally seen as “Junk DNA”. These findings have been interpreted as fundamentally undermining the “Junk DNA” concept. Here, we reinforce previous arguments that this interpretation relies on an inadequate concept of biological function that does not consider the selected effect of a given genomic structure, which is central to the “Junk DNA” concept. Next, we suggest that another (though ignored) confounding factor is that the discussion about biological functions includes two different dimensions: a horizontal, ecological dimension that reflects how a given genomic element affects fitness in a specific time, and a vertical, temporal dimension that reflects how a given genomic element persisted along time. We suggest that “Junk DNA” should be used exclusively relative to the horizontal dimension, while for the vertical dimension, we propose a new term, “Spam DNA”, that reflects the fact that a given genomic element may persist in the genome even if not selected for on their origin. Importantly, these concepts are complementary. An element can be both “Spam DNA” and “Junk DNA”, and “Spam DNA” can also be recruited to perform evolved biological functions, as illustrated in processes of exaptation or constructive neutral evolution.

https://doi.org/10.1093/gbe/evac055 - 2022

PubReading [308] - Deep learning in cancer pathology- a new generation of clinical biomarkers - A. Echle, J. Kather et al.

Thu, 16 Mar 2023 12:00:26 GMT

Clinical workflows in oncology rely on predictive and prognostic molecular biomarkers. However, the growing number of these complex biomarkers tends to increase the cost and time for decision-making in routine daily oncology practice; furthermore, biomarkers often require tumour tissue on top of routine diagnostic material. Nevertheless, routinely available tumour tissue contains an abundance of clinically relevant information that is currently not fully exploited. Advances in deep learning (DL), an artificial intelligence (AI) technology, have enabled the extraction of previously hidden information directly from routine histology images of cancer, providing potentially clinically useful information. Here, we outline emerging concepts of how DL can extract biomarkers directly from histology images and summarise studies of basic and advanced image analysis for cancer histology. Basic image analysis tasks include detection, grading and subtyping of tumour tissue in histology images; they are aimed at automating pathology workflows and consequently do not immediately translate into clinical decisions. Exceeding such basic approaches, DL has also been used for advanced image analysis tasks, which have the potential of directly affecting clinical decision-making processes. These advanced approaches include inference of molecular features, prediction of survival and end-to-end prediction of therapy response. Predictions made by such DL systems could simplify and enrich clinical decision-making, but require rigorous external validation in clinical settings.

https://doi.org/10.1038/s41416-020-01122-x - 2020

PubReading [307] - Comparison of X-ray and NMR Structures- Is There a Systematic Difference in Residue Contacts between X-ray- and NMR-Resolved Protein Structures? - S. Garbuzynskiy, O. Galzitskaya et al.

Wed, 15 Mar 2023 12:00:32 GMT

We have compared structures of 78 proteins determined by both NMR and X-ray methods. It is shown that X-ray and NMR structures of the same protein have more differences than various X-ray structures obtained for the protein, and even more than various NMR structures of the protein. X-ray and NMR structures of 18 of these 78 proteins have obvious large-scale structural differences that seem to reflect a difference of crystal and solution structures. The other 60 pairs of structures have only small-scale differences comparable with differences between various X-ray or various NMR structures of a protein; we have analyzed these structures more attentively. One of the main differences between NMR and X-ray structures concerns the number of contacts per residue: NMR structures presented in PDB have more contacts than X-ray structures at distances below 3.0 Å and 4.5– 6.5 Å, and fewer contacts at distances of 3.0 – 4.5 Å and 6.5–8.0 Å; this difference in the number of contacts is greater for internal residues than for external ones, and it is larger for -containing proteins than for all proteins. Another significant difference is that the main-chain hydrogen bonds identified in X-ray and NMR structures often differ. Their correlation is 69% only. However, analogous difference is found for refined and rerefined NMR structures, allowing us to suggest that the observed difference in interresidue contacts of X-ray and NMR structures of the same proteins is due mainly to a difference in mathematical treatment of experimental results.

DOI: 10.1002/prot.20491 - 2005

PubReading [306] - A Brief History of IL-9 - R. Goswami and M. Kaplan

Tue, 14 Mar 2023 12:00:20 GMT

IL-9 was first described in the late 1980s as a member of a growing number of cytokines that had pleiotropic functions in the immune system. Although many biological functions have been attributed to IL-9, it remains an understudied cytokine. A resurgence of interest in IL-9 has been spurred by recent work demonstrating a role for IL-9 in regulating inflammatory immunity and defining the transcription factors that activate the Il9 gene in cells that most efficiently produce IL-9. In this review, we summarize the characterization of IL-9 biological activities, highlight roles for the cytokine that are clearly defined, and outline questions regarding IL-9 functions that still require further exploration.

www.jimmunol.org/cgi/doi/10.4049/jimmunol.1003049 - 2011

PubReading [304] - What are Biomarkers? - K. Strimbu and J. Tavel

Fri, 10 Mar 2023 12:00:36 GMT

This article provides working definitions and a conceptual framework to understand the roles of biomarkers in clinical research. The definitions of the terms discussed in this article—medical signs, symptoms, biomarkers, surrogate endpoints, clinical endpoints, validation—are still under discussion, as are their relationships to each other, but broad consensus has developed in the past decade and a half about the necessity of distinguishing between, in particular, surrogate and clinical endpoints.

doi:10.1097/COH.0b013e32833ed177. - 2010

PubReading [303] - The Epigenomics of Cancer - P. Jones and S. Baylin

Thu, 09 Mar 2023 12:00:27 GMT

Aberrant gene function and altered patterns of gene expression are key features of cancer. Growing evidence shows that acquired epigenetic abnormalities participate with genetic alterations to cause this dysregulation. Here, we review recent advances in understanding how epigenetic alterations participate in the earliest stages of neoplasia, including stem/ precursor cell contributions, and discuss the growing implications of these advances for strategies to control cancer.

DOI 10.1016/j.cell.2007.01.029 -2007

PubReading [302] - The evolution, evolvability and engineering of gene regulatory DNA - E. Vaishnav, A. Regev et al.

Wed, 08 Mar 2023 12:00:18 GMT

Mutations in non-coding regulatory DNA sequences can alter gene expression, organismal phenotype and fitness1–3. Constructing complete fitness landscapes, in which DNA sequences are mapped to fitness, is a long-standing goal in biology, but has remained elusive because it is challenging to generalize reliably to vast sequence spaces4–6. Here we build sequence-to-expression models that capture fitness landscapes and use them to decipher principles of regulatory evolution. Using millions of randomly sampled promoter DNA sequences and their measured expression levels in the yeast Saccharomyces cerevisiae, we learn deep neural network models that generalize with excellent prediction performance, and enable sequence design for expression engineering. Using our models, we study expression divergence under genetic drift and strong-selection weak-mutation regimes to find that regulatory evolution is rapid and subject to diminishing returns epistasis; that conflicting expression objectives in different environments constrain expression adaptation; and that stabilizing selection on gene expression leads to the moderation of regulatory complexity. We present an approach for using such models to detect signatures of selection on expression from natural variation in regulatory sequences and use it to discover an instance of convergent regulatory evolution. We assess mutational robustness, finding that regulatory mutation effect sizes follow a power law, characterize regulatory evolvability, visualize promoter fitness landscapes, discover evolvability archetypes and illustrate the mutational robustness of natural regulatory sequence populations. Our work provides a general framework for designing regulatory sequences and addressing fundamental questions in regulatory evolution.

https://doi.org/10.1038/s41586-022-04506-6 - 2022

PubReading [301] - Cancer Progress and Priorities- Lung Cancer - M. Schabath and M. Cote

Tue, 07 Mar 2023 12:01:01 GMT

In the United States, lung cancer is the second most common diagnosed cancer and the leading cause of cancer-related death. Although tobacco smoking is the major risk factor accounting for 80% to 90% of all lung cancer diagnoses, there are numerous other risk factors that have been identified as casually associated with lung cancer etiology. However, there are few causally linked risk factors for lung cancer diagnosed among never smokers, which, if considered a unique reportable category, is the 11th most common cancer and the 7th leading cause of cancer-related death. Lung cancer survival has only marginally improved over the last several decades, but the availability of screening and early detection by low-dose CT and advances in targeted treatments and immunotherapy will likely decrease mortality rates and improve patient survival outcomes in the near future.

doi: 10.1158/1055-9965.EPI-19-0221 - 2019

PubReading [299] - The Formation and Displacement of Ordered DNA Triplexes in Self- Assembled Three-Dimensional DNA Crystals - Y. Zhao, R. Sha et al.

Fri, 03 Mar 2023 12:00:34 GMT

Reconfigurable structures engineered through DNA hybridization and self-assembly offer both structural and dynamic applications in nanotechnology. Here, we have demonstrated that strand displacement of triplex-forming oligonucleotides (TFOs) can be translated to a robust macroscopic DNA crystal by coloring the crystals with covalently attached fluorescent dyes. We show that three different types of triplex strand displacement are feasible within the DNA crystals and the bound TFOs can be removed and/or replaced by (a) changing the pH from 5 to 7, (b) the addition of the Watson–Crick complement to a TFO containing a short toehold, and (c) the addition of a longer TFO that uses the duplex edge as a toehold. We have also proved by X-ray diffraction that the structure of the crystals remains as designed in the presence of the TFOs.

https://doi.org/10.1021/jacs.2c12667 - 2023

PubReading [298] - Simultaneous sequencing of genetic and epigenetic bases in DNA - J. Füllgrabe, S. Balasubramanian et al.

Thu, 02 Mar 2023 12:00:38 GMT

DNA comprises molecular information stored in genetic and epigenetic bases, both of which are vital to our understanding of biology. Most DNA sequencing approaches address either genetics or epigenetics and thus capture incomplete information. Methods widely used to detect epigenetic DNA bases fail to capture common C-to-T mutations or distinguish 5-methylcytosine from 5-hydroxymethylcytosine. We present a single base-resolution sequencing methodology that sequences complete genetics and the two most common cytosine modifications in a single workflow. DNA is copied and bases are enzymatically converted. Coupled decoding of bases across the original and copy strand provides a phased digital readout. Methods are demonstrated on human genomic DNA and cell-free DNA from a blood sample of a patient with cancer. The approach is accurate, requires low DNA input and has a simple workflow and analysis pipeline. Simultaneous, phased reading of genetic and epigenetic bases provides a more complete picture of the information stored in genomes and has applications throughout biomedicine.

https://doi.org/10.1038/s41587-022-01652-0 - 2022

PubReading [297] - Modulation of the helical properties of DNA- next-to-nearest neighbour effects and beyond - A. Balaceanu, M. Orozco

Wed, 01 Mar 2023 12:00:20 GMT

We used extensive molecular dynamics simulations to study the structural and dynamic properties of the central d(TpA) step in the highly polymorphic d(CpTpApG) tetranucleotide. Contrary to the assumption of the dinucleotide model and its nearest neighbours (tetranucleotide-model), the properties of the central d(TpA) step change quite significantly dependent on the next-to-nearest (hexanucleotide) sequence context and in a few cases are modulated by even remote neighbours (beyond next-to-nearest from the central TpA). Our results highlight the existence of previously undescribed dynamical mechanisms for the transmission of structural information into the DNA and demonstrate the existence of certain sequences with special physical properties that can impact on the global DNA structure and dynamics.

PubReading [296] - Telomerase structural biology comes of age - Y. He & J. Feigon

Tue, 28 Feb 2023 12:00:39 GMT

Telomerase is an RNA–protein complex comprising telomerase reverse transcriptase, a non-coding telomerase RNA, and proteins involved in biogenesis, assembly, localization, or recruitment. Telomerase synthesizes the telomeric DNA at the 30-ends of linear chromosomes. During the past decade, structural studies have defined the architecture of Tetrahymena and human telomerase as well as protein and RNA domain structures, but high-resolution details of interactions remained largely elusive. In the past two years, several sub-4 Å cryo-electron microscopy structures of telomerase were published, including Tetrahymena telomerase at different steps of telomere repeat addition and human telomerase with telomere shelterin proteins that recruit telomerase to telomeres. These and other recent structural studies have expanded our understanding of telomerase assembly, mechanism, recruitment, and mutations leading to disease.

https://doi.org/10.1016/j.sbi.2022.102446 - 2022

PubReading [294] - Violation of DNA neighbor exclusion principle in RNA recognition - M. Yousuf, K. Kim et al.

Fri, 24 Feb 2023 12:00:24 GMT

DNA intercalation has been very useful for engineering DNA-based functional materials. It is generally expected that the intercalation phenomenon in RNA would be similar to that in DNA. Here we note that the neighbor-exclusion principle is violated in RNA by naphthalene-based cationic probes, in contrast to the fact that it is usually valid in DNA. All the intercalation structures are responsible for the fluorescence, where small naphthalene moieties are intercalated in between bases via p–p interactions. The structure is aided by hydrogen bonds between the cationic moieties and the ribose-phosphate backbone, which results in specific selectivity for RNA over DNA. This experimentally observed mechanism is supported by computationally reproducing the fluorescence and CD data. MD simulations confirm the unfolding of RNA due to the intercalation of probes. Elucidation of the mechanism of selective sensing for RNA over DNA would be highly beneficial for dynamical observation of RNA which is essential for studying its biological roles.

DOI: 10.1039/c5sc03740a - 2016

PubReading [293] - The Nuclear Factor Kappa B (NF-kB) signaling in cancer development and immune diseases - M. Zinatizadeh, S. Miri et al.

Thu, 23 Feb 2023 13:00:17 GMT

The nuclear factor kappa B (NF-kB) family of transcription factors plays an essential role as stressors in the cellular environment, and controls the expression of important regulatory genes such as immunity, inflammation, death, and cell proliferation. NF-kB protein is located in the cytoplasm, and can be activated by various cellular stimuli. There are two pathways for NF-kB activation, as the canonical and non-canonical pathways, which require complex molecular interactions with adapter proteins and phosphorylation and ubiquitinase enzymes. Accordingly, this increases NF-kB translocation in the nucleus and regulates gene expression. In this study, the concepts that emerge in different cellular systems allow the design of NF-kB function in humans. This would not only allow the development for rare diseases associated with NF-kB, but would also be used as a source of useful information to eliminate widespread consequences such as cancer or inflammatory/immune diseases.

https://doi.org/10.1016/j.gendis.2020.06.005 - 2020

PubReading [292] - Acquisition of Dynamic Function in Human Stem Cell-Derived b Cells - L. Velazco-Cruz, J. Millman et al.

Wed, 22 Feb 2023 14:00:45 GMT

Recent advances in human pluripotent stem cell (hPSC) differentiation protocols have generated insulin-producing cells resembling pancreatic b cells. While these stem cell-derived b (SC-b) cells are capable of undergoing glucose-stimulated insulin secretion (GSIS), insulin secretion per cell remains low compared with islets and cells lack dynamic insulin release. Herein, we report a differentiation strategy focused on modulating transforming growth factor b (TGF-b) signaling, controlling cellular cluster size, and using an enriched serum-free media to generate SC-b cells that express b cell markers and undergo GSIS with first- and second-phase dynamic insulin secretion. Transplantation of these cells into mice greatly improves glucose tolerance. These results reveal that specific time frames for inhibiting and permitting TGF-b signaling are required during SC-b cell differentiation to achieve dynamic function. The capacity of these cells to undergo GSIS with dynamic insulin release makes them a promising cell source for diabetes cellular therapy.

https://doi.org/10.1016/j.stemcr.2018.12.012 - 2019

PubReading [291] - Tumour Cell Secretome in Chemoresistance and Tumour Recurrence - E. Madden, A. Samali et al.

Tue, 21 Feb 2023 13:00:27 GMT

Chemoresistance is a major factor driving tumour relapse and the high rates of cancer-related deaths. Understanding how cancer cells overcome chemotherapy-induced cell death is critical in promoting patient survival. One emerging mechanism of chemoresistance is the tumour cell secretome (TCS), an array of protumorigenic factors released by tumour cells. Chemotherapy exposure can also alter the composition of the TCS, known as therapy-induced TCS, and can promote tumour relapse and the formation of an immunosuppressive tumour microenvironment (TME). Here, we outline how the TCS can protect cancer cells from chemotherapy-induced cell death. We also highlight recent evidence describing how therapy-induced TCS can impact cancer stem cell (CSC) expansion and tumour-associated immune cells to enable tumour regrowth and antitumour immunity.

https://doi.org/10.1016/j.trecan.2020.02.020 - 2020

PubReading [289] - The importance of loop length on the stability of i-motif structures - S. Gurung, C. Schwarz, J. Hall, C. Cardin and J. Brazier

Fri, 17 Feb 2023 12:00:19 GMT

Using UV and srCD spectroscopy it is found that loop length within the i-motif structure is important for both thermal and pH stability, but in contrast to previous statements, it is the shorter loops that exhibit the highest stability.

DOI: 10.1039/c4cc07279k - 2015

PubReading [288] - Structural and functional diversity calls for a new classification of ABC transporters - C. Thomas, J. Zimmer and R. Tampe

Thu, 16 Feb 2023 13:00:18 GMT

Members of the ATP-binding cassette (ABC) transporter superfamily translocate a broad spectrum of chemically diverse substrates. While their eponymous ATP-binding cassette in the nucleotide-binding domains (NBDs) is highly conserved, their transmembrane domains (TMDs) forming the translocation pathway exhibit distinct folds and topologies, suggesting that during evolution the ancient motor domains were combined with different transmembrane mechanical systems to orchestrate a variety of cellular processes. In recent years, it has become increasingly evident that the distinct TMD folds are best suited to categorize the multitude of ABC transporters. We therefore propose a new ABC transporter classification that is based on structural homology in the TMDs.

doi:10.1002/1873-3468.13935 - 2020

PubReading [287] - On the Origin of the Term ‘‘Stem Cell’’ - M. Ramalho-Santos and H. Willenbring

Wed, 15 Feb 2023 13:00:17 GMT

Stem cells have fascinated both biologists and clinicians for over a century. Here, we discuss the origin of the term ‘‘stem cell,’’ which can be traced back to the late 19th century. The term stem cell originated in the context of two major embryological questions of that time: the continuity of the germ-plasm and the origin of the hematopoietic system. Theodor Boveri and Valentin Hacker used the term stem cell to describe cells committed to give rise to the germline. In parallel, Artur Pappenheim, Alexander Maximow, Ernst Neumann, and others used it to describe a proposed progenitor of the blood system. The original meanings of the term stem cell, rather than being historical relics, continue to capture important aspects of the biology of stem cells as we see them today.

DOI 10.1016/j.stem.2007.05.013 - 2007

PubReading [286] - Cancer occurrence in Turner syndrome and the effect of sex hormone substitution therapy - M. Viuff, C. Gravholt et al.

Tue, 14 Feb 2023 13:00:17 GMT

Although the overall risk of cancer is not increased in Turner syndrome, the pattern of cancer occurrence differs from the general population. We aim to describe the cancer morbidity pattern in Turner syndrome and evaluate the effect of long-term hormone replacement therapy (HRT). Design: Nationwide epidemiological study. The lack of one X chromosome might play a role in skin neoplasms, CNS tumors, colon and rectal cancers. The risk of breast cancer is lower than in the general population. Long-term HRT during the premenopausal age range seems not to exert a cancerous effect in Turner syndrome. Increased vigilance concerning specific types of cancer in Tuner syndrome harboring a 45,X karyotype is needed.

https://doi.org/10.1530/EJE-20-0702 - 2021

PubReading [284] - A brief history of nearly EV-erything – The rise and rise of extracellular vesicles - Y. Couch, D. Carter et al.

Fri, 10 Feb 2023 12:00:23 GMT

Extracellular vesicles (EVs) are small cargo-bearing vesicles released by cells into the extracellular space. The field of EVs has grown exponentially over the past two decades; this growth follows the realisation that EVs are not simply a waste disposal system as had originally been suggested by some, but also a complex cell-to-cell communication mechanism. Indeed, EVs have been shown to transfer functional cargo between cells and can influence several biological processes. These small biological particles are also deregulated in disease. As we approach the 75th anniversary of the first experiments in which EVs were unknowingly isolated, it seems right to take stock and look back on how the field started, and has since exploded into its current state. Here we review the early experiments, summarise key findings that have propelled the field, describe the growth of an organised EV community, discuss the current state of the field, and identify key challenges that need to be addressed.

DOI: 10.1002/jev2.12144 - 2021

PubReading [283] - PARP1 Gene Knockout Suppresses Expression of DNA Base Excision Repair Genes - A. Zakharenkoa, O. Lavrik et al.

Thu, 09 Feb 2023 13:00:27 GMT

The effect of PARP1 knockout in HEK293 cells on the gene expression of DNA base excision repair (BER) proteins was studied. It was shown that the expression of all differentially expressed genes (DEGs) of BER was reduced by knockout. The expression of the DNA glycosylase gene NEIL1, which is considered to be one of the common “hubs” for binding BER proteins, has changed the most. The expression of genes of auxiliary subunits of DNA polymerases δ and ε is also significantly reduced. The PARP1 gene knock-out cell line obtained is an adequate cell model for studying the activity of the BER process in the absence of PARP1 and testing drugs aimed at inhibiting repair processes. It has been found for the first time that knock-out of the PARP1 gene results in a significant change in the level of expression of proteins responsible for ribosome biogenesis and the functioning of the proteasome.

DOI: 10.1134/S1607672922700028 - 2022

PubReading [282] - Modified RNA triplexes- Thermodynamics, structure and biological potential - M. Szabat, E. Kierzek & R. Kierzek

Wed, 08 Feb 2023 13:00:15 GMT

The occurrence of triplexes in vivo has been well documented and is determined by the presence of long homopurine-homopyrimidine tracts. The formation of these structures is the result of conformational changes that occur in the duplex, which allow the binding of a third strand within the major groove of the helix. Formation of these noncanonical forms by introducing synthetic triplex-forming oligonucleotides (TFOs) into the cell may have applications in molecular biology, diagnostics and therapy. This study focused on the formation of RNA triplexes as well as their thermal stability and biological potential in the HeLa cell line. Thermodynamics studies revealed that the incorporation of multiple locked nucleic acid (LNA) and 2-thiouridine (2-thioU) residues increased the stability of RNA triplexes. These data suggest that the number and position of the modified nucleotides within TFOs significantly stabilize the formed structures. Moreover, specificity of the interactions between the modified TFOs and the RNA hairpin was characterized using electrophoretic mobility-shift assay (EMSA), and triplex dissociation constants have been also determined. Finally, through quantitative analysis of GFP expression, the triplex structures were shown to regulate GFP gene silencing. Together, our data provide a first glimpse into the thermodynamic, structural and biological properties of LNA- and 2-thioU modified RNA triplexes.

DOI:10.1038/s41598-018-31387-5 - 2018

PubReading [281] - Light-Responsive Dynamic DNA-Origami-Based Plasmonic Assemblies - J. Ryssy, A. Kuzyk et al.

Tue, 07 Feb 2023 13:00:15 GMT

DNA nanotechnology offers a versatile toolbox for precise spatial and temporal manipulation of matter on the nanoscale. However, rendering DNA-based systems responsive to light has remained challenging. Herein, we describe the remote manipulation of native (non-photoresponsive) chiral plasmonic molecules (CPMs) using light. Our strategy is based on the use of a photoresponsive medium comprising a merocyanine-based photoacid. Upon exposure to visible light, the medium decreases its pH, inducing the formation of DNA triplex links, leading to a spatial reconfiguration of the CPMs. The process can be reversed simply by turning the light off and it can be repeated for multiple cycles. The degree of the overall chirality change in an ensemble of CPMs depends on the CPM fraction undergoing reconfiguration, which, remarkably, depends on and can be tuned by the intensity of incident light. Such a dynamic, remotely controlled system could aid in further advancing DNA-based devices and nanomaterials.

doi.org/10.1002/anie.202014963 - 2021

PubReading [279] - Facts and Hopes in Immunotherapy for Early-Stage Triple-Negative Breast Cancer - I. Nederlof, L. Voorwerk & M. Kok

Fri, 03 Feb 2023 13:00:19 GMT

A substantial fraction of early-stage triple-negative breast cancer (eTNBC) is characterized by high levels of stromal tumor-infiltrating lymphocytes (sTIL) and has a good prognosis even without systemic treatment, highlighting the importance of an endogenous anticancer immune response. Still, a considerable proportion of patients with eTNBC need some “therapeutical push” to kick-start this immune response. Exploiting this immune response with immune-checkpoint inhibition (ICI), in combination with chemotherapy, has made its way into standard of care in eTNBC. Major challenges in the near future include finding those patients with eTNBC who can be treated with ICI alone or with a reduced chemotherapy backbone. Exploring the optimal duration of ICI and finding biomarkers to predict response will be key to enable personalized implementation of ICI in patients with eTNBC. For patients who currently do not respond effectively to ICI plus chemotherapy, challenges lie in finding new immunomodulatory therapies and developing response-guided neoadjuvant approaches.

doi: 10.1158/1078-0432.CCR-22-0701 - 2023

PubReading [278] - Light-Enhanced Cytotoxicity of Doxorubicin by Photoactivation - G. Greco, M. Calvaresi et al.

Thu, 02 Feb 2023 13:00:46 GMT

The combination of photodynamic therapy with chemotherapy (photochemotherapy, PCT) can lead to additive or synergistic antitumor effects. Usually, two different molecules, a photosensitizer (PS) and a chemotherapeutic drug are used in PCT. Doxorubicin is one of the most successful chemotherapy drugs. Despite its high efficacy, two factors limit its clinical use: severe side effects and the development of chemoresistance. Doxorubicin is a chromophore, able to absorb light in the visible range, making it a potential PS. Here, we exploited the intrinsic photosensitizing properties of doxorubicin to enhance its anticancer activity in leukemia, breast, and epidermoid carcinoma cells, upon irradiation. Light can selectively trigger the local generation of reactive oxygen species (ROS), following photophysical pathways. Doxorubicin showed a concentration-dependent ability to generate peroxides and singlet oxygen upon irradiation. The underlying mechanisms leading to the increase in its cytotoxic activity were intracellular ROS generation and the induction of necrotic cell death. The nuclear localization of doxorubicin represents an added value for its use as a PS. The use of doxorubicin in PCT, simultaneously acting as a chemotherapeutic agent and a PS, may allow (i) an increase in the anticancer effects of the drug, and (ii) a decrease in its dose, and thus, its dose-related adverse effects.

https://doi.org/ 10.3390/cells12030392 - 2023

PubReading [277] - Direct targeting of amplified gene loci for proapoptotic anticancer therapy - M. Tiwari, F. Rogers at al..

Wed, 01 Feb 2023 13:00:33 GMT

Gene amplification drives oncogenesis in a broad spectrum of cancers. A number of drugs have been developed to inhibit the protein products of amplified driver genes, but their clinical efficacy is often hampered by drug resistance. Here, we introduce a therapeutic strategy for targeting cancer-associated gene amplifications by activating the DNA damage response with triplex-forming oligonucleotides (TFOs), which drive the induction of apoptosis in tumors, whereas cells without amplifications process lower levels of DNA damage. Focusing on cancers driven by HER2 amplification, we find that TFOs targeting HER2 induce copy number-dependent DNA double-strand breaks (DSBs) and activate p53-independent apoptosis in HER2-positive cancer cells and human tumor xenografts via a mechanism that is independent of HER2 cellular function. This strategy has demonstrated in vivo efficacy comparable to that of current precision medicines and provided a feasible alternative to combat drug resistance in HER2-positive breast and ovarian cancer models. These findings offer a general strategy for targeting tumors with amplified genomic loci.

https://doi.org/10.1038/s41587-021-01057-5 - 2022

PubReading [276] - Impact of 3D cell culture on bone regeneration potential of mesenchymal stromal cells - M. Bicer, G. Cottrell and D. Widera

Tue, 31 Jan 2023 13:00:28 GMT

As populations age across the world, osteoporosis and osteoporosis-related fractures are becoming the most prevalent degenerative bone diseases. More than 75 million patients suffer from osteoporosis in the USA, the EU and Japan. Furthermore, it is anticipated that the number of patients affected by osteoporosis will increase by a third by 2050. Although conventional therapies including bisphosphonates, calcitonin and oestrogen-like drugs can be used to treat degenerative diseases of the bone, they are often associated with serious side effects including the development of oesophageal cancer, ocular inflammation, severe musculoskeletal pain and osteonecrosis of the jaw. The use of autologous mesenchymal stromal cells/mesenchymal stem cells (MSCs) is a possible alternative therapeutic approach to tackle osteoporosis while overcoming the limitations of traditional treatment options. However, osteoporosis can cause a decrease in the numbers of MSCs, induce their senescence and lower their osteogenic differentiation potential. Three-dimensional (3D) cell culture is an emerging technology that allows a more physiological expansion and differentiation of stem cells compared to cultivation on conventional flat systems. This review will discuss current understanding of the effects of different 3D cell culture systems on proliferation, viability and osteogenic differentiation, as well as on the immunomodulatory and anti-inflammatory potential of MSCs.

https://doi.org/10.1186/s13287-020-02094-8 - 2021

PubReading [274] - The eukaryotic replisome tolerates leading-strand base damage by replicase switching - T. Guilliam & J. Yeeles

Fri, 27 Jan 2023 12:00:16 GMT

The high-fidelity replicative DNA polymerases, Pol e and Pol d, are generally thought to be poorly equipped to replicate damaged DNA. Direct and complete replication of a damaged template therefore typically requires the activity of low-fidelity translesion synthesis (TLS) polymerases. Here we show that a yeast replisome, reconstituted with purified proteins, is inherently tolerant of the common oxidative lesion thymine glycol (Tg). Surprisingly, leading-strand Tg was bypassed efficiently in the presence and absence of the TLS machinery. Our data reveal that following helicase–polymerase uncoupling a switch from Pol e, the canonical leading-strand replicase, to the lagging-strand replicase Pol d, facilitates rapid, efficient and error-free lesion bypass at physiological nucleotide levels. This replicase switch mechanism also promotes bypass of the unrelated oxidative lesion, 8-oxoguanine. We propose that replicase switching may promote continued leading-strand synthesis whenever the replisome encounters leading-strand damage that is bypassed more efficiently by Pol d than by Pol e.

DOI: 10.15252/embj.2020107037 - 20121

PubReading [273] - RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2 - J. Bravo, D. Taylor et al.

Thu, 26 Jan 2023 12:00:15 GMT

Cas12a2 is a CRISPR-associated nuclease that performs RNA-guided, sequence- nonspecific degradation of single-stranded RNA, single-stranded DNA and double- stranded DNA following recognition of a complementary RNA target, culminating in abortive infection1. Here we report structures of Cas12a2 in binary, ternary and quaternary complexes to reveal a complete activation pathway. Our structures reveal that Cas12a2 is autoinhibited until binding a cognate RNA target, which exposes the RuvC active site within a large, positively charged cleft. Double-stranded DNA substrates are captured through duplex distortion and local melting, stabilized by pairs of ‘aromatic clamp’ residues that are crucial for double-stranded DNA degradation and in vivo immune system function. Our work provides a structural basis for this mechanism of abortive infection to achieve population-level immunity, which can be leveraged to create rational mutants that degrade a spectrum of collateral substrates.

https://doi.org/10.1038/s41586-022-05560-w - 2022

PubReading [272] - Frailty biomarkers under the perspective of geroscience- A narrative review - R. Dos Santos Aguiar Gonçalves, P. de Souto Barreto

Wed, 25 Jan 2023 12:00:40 GMT

Cellular and molecular aging biomarkers might contribute to identify at-risk individuals for frailty before overt clinical manifestations appear. Although studies on the associations of aging biomarkers and frailty exist, no investigation has gathered this information using a structured framework for identifying aging biomarkers; as a result, the evidence on frailty and aging biomarkers is diffuse and incomplete. Therefore, this narrative review aimed to gather information on the associations of the hallmarks of aging and frailty under the perspective of geroscience. The literature on human studies on this topic is sparse and mainly composed of cross-sectional investigations performed in small study samples. The main putative aging biomarkers associated to frailty were: mitochondrial DNA copy number (genomic instability and mitochondrial dysfunction), telomere length (telomere attrition), global DNA methylation (epigenetic alterations), Hsp70 and Hsp72 (loss of proteostasis), IGF-1 and SIRT1 (deregulated nutrient-sensing), GDF-15 (mitochondrial dysfunction, cellular senescence and altered intercellular communication), CD4 + and CD8 + cell percentages (cellular senescence), circulating osteogenic progenitor (COP) cells (stem cell exhaustion), and IL-6, CRP and TNF-alpha (altered intercellular communication). IGF-1, SIRT1, GDF-15, IL-6, CRP and TNF-alpha presented more evidence among these biomarkers, highlighting the importance of inflammation and nutrient sensing on frailty. Further longitudinal studies investigating biomarkers across the hallmarks of aging would provide valuable information on this topic.

https://doi.org/10.1016/j.arr.2022.101737 - 2022

PubReading [271] - Mass-resolved electronic circular dichroism ion spectroscopy - S. Daly, F. Rosu, V. Gabelica

Tue, 24 Jan 2023 12:00:19 GMT

DNA and proteins are chiral: Their three-dimensional structures cannot be superimposed with their mirror images. Circular dichroism spectroscopy is widely used to characterize chiral compounds, but data interpretation is difficult in the case of mixtures. We recorded the electronic circular dichroism spectra of DNA helices separated in a mass spectrometer. We studied guanine-rich strands having various secondary structures, electrosprayed them as negative ions, irradiated them with an ultraviolet nanosecond optical parametric oscillator laser, and measured the difference in electron photodetachment efficiency between left and right circularly polarized light. The reconstructed circular dichroism ion spectra resembled those of their solution-phase counterparts, thereby allowing us to assign the DNA helical topology. The ability to measure circular dichroism directly on biomolecular ions expands the capabilities of mass spectrometry for structural analysis.

DOI: 10.1126/science.abb1822 - 2020

PubReading [269] - Rapid and reversible optogenetic silencing of synaptic transmission by clustering of synaptic vesicles - D. Vettkötter, A. Gottschalk et al.

Fri, 20 Jan 2023 12:00:15 GMT

Acutely silencing specific neurons informs about their functional roles in circuits and behavior. Existing optogenetic silencers include ion pumps, channels, metabotropic receptors, and tools that damage the neurotransmitter-release machinery. While the former hyperpolarize the cell, alter ionic gradients or cellular biochemistry, the latter allow only slow recovery, requiring de novo synthesis. Thus, tools combining fast activation and reversibility are needed. Here, we use light-evoked homo-oligomerization of cryptochrome CRY2 to silence synaptic transmission, by clustering synaptic vesicles (SVs). We benchmark this tool, optoSynC, in Caenorhabditis elegans, zebrafish, and murine hippocampal neurons. optoSynC clusters SVs, observable by electron microscopy. Locomotion silencing occurs with tauon ~7.2 s and recovers with tauoff ~6.5 min after light-off. optoSynC can inhibit exocytosis for several hours, at very low light intensities, does not affect ion currents, biochemistry or synaptic proteins, and may further allow manipulating different SV pools and the transfer of SVs between them.

https://doi.org/10.1038/s41467-022-35324-z - 2022

PubReading [268] - Inheritance of paternal DNA damage by histone-mediated repair restriction - S. Wang, D. Meyer & B. Schumacher

Thu, 19 Jan 2023 12:00:16 GMT

How paternal exposure to ionizing radiation affects genetic inheritance and disease risk in the offspring has been a long-standing question in radiation biology. In humans, nearly 80% of transmitted mutations arise in the paternal germline1, but the transgenerational effects of ionizing radiation exposure has remained controversial and the mechanisms are unknown. Here we show that in sex-separated Caenorhabditis elegans strains, paternal, but not maternal, exposure to ionizing radiation leads to transgenerational embryonic lethality. The offspring of irradiated males displayed various genome instability phenotypes, including DNA fragmentation, chromosomal rearrangement and aneuploidy. Paternal DNA double strand breaks were repaired by maternally provided error-prone polymerase theta-mediated end joining. Mechanistically, we show that depletion of an orthologue of human histone H1.0, HIS-24, or the heterochromatin protein HPL-1, could significantly reverse the transgenerational embryonic lethality. Removal of HIS-24 or HPL-1 reduced histone 3 lysine 9 dimethylation and enabled error-free homologous recombination repair in the germline of the F1 generation from ionizing radiation-treated P0 males, consequently improving the viability of the F2 generation. This work establishes the mechanistic underpinnings of the heritable consequences of paternal radiation exposure on the health of offspring, which may lead to congenital disorders and cancer in humans.

https://doi.org/10.1038/s41586-022-05544-w - 2022

PubReading [267] - Circular dichroism and its uses in biomolecular research - A Review - Sangeeta and Anu Radha Pathania

Wed, 18 Jan 2023 12:00:33 GMT

The higher-order structure of proteins as well as their thermal stability can be determined using the circular dichroism (CD). CD is a common approach for swiftly assessing binding, secondary structure, and folding properties of proteins. In a nutshell, circular dichroism is an absorption spectroscopy technique that employs circularly polarized light to explore structural properties of optically active chiral compounds. Biological molecules, as well as their interactions with metals and other compounds, are studied extensively. Circular dichroism is becoming more widely acknowledged as a useful technique for studying the various conformations taken by proteins and nucleic acids in solution. Because CD is a quantitative approach, it can be used to track protein denaturation and protein-ligand interaction. These CD measures will have two key advantages: they can be performed on small amounts of material in a physiological buffer, and they will provide one of the greatest methods for monitoring any structural changes that occur as a result of changes in environmental conditions. It has proven possible to generate proteins on a big scale for therapeutic reasons utilizing recombinant DNA technology. Circular dichroism is also well-known as a useful method which is used for determining the folding characteristics of proteins. CD is used to see if a purified, produced peptide is either bended or if it has a mutation that impacts its strength and confirmation. The basic steps in getting this CD data, as well as the methodologies for interpreting the spectra in order to predict the protein structure, are summarized in this article. However, many researchers' value is harmed when they use circular dichroism, either because of poor experimental design or because of insufficient data. The essential steps in getting this CD data, as well as the methodologies for interpreting the spectra in order to predict the protein structure, will be summarized in this article. However, the value of many investigations using circular dichroism is harmed due to insufficient attention to critical components of instrument calibration or sample characterization.

https://doi.org/10.1051/e3sconf/202130901095 - 2021

PubReading [266] - High-pressure crystallography shows noble gas intervention into protein-lipid interaction and suggests a model for anaesthetic action - I. Melnikov, V. Gordeliy, A. Popov et al.

Tue, 17 Jan 2023 13:00:30 GMT

In this work we examine how small hydrophobic molecules such as inert gases interact with membrane proteins (MPs) at a molecular level. High pressure atmospheres of argon and krypton were used to produce noble gas derivatives of crystals of three well studied MPs (two different proton pumps and a sodium light-driven ion pump). The structures obtained using X-ray crystallography showed that the vast majority of argon and krypton binding sites were located on the outer hydrophobic surface of the MPs – a surface usually accommodating hydrophobic chains of annular lipids (which are known structural and functional determinants for MPs). In conformity with these results, supplementary in silico molecular dynamics (MD) analysis predicted even greater numbers of argon and krypton binding positions on MP surface within the bilayer. These results indicate a potential importance of such interactions, particularly as related to the phenomenon of noble gas-induced anaesthesia.

https://doi.org/10.1038/s42003-022-03233-y - 2022

PubReading [264] - Role of astrocytes in rhythmic motor activity - A. Montalant, E. Carlsen, JF Perrier

Fri, 13 Jan 2023 13:00:19 GMT

Rhythmic motor activities such as breathing, locomotion, tremor, or mastication are organized by groups of interconnected neurons. Most synapses in the central nervous system are in close apposition with processes belonging to astrocytes. Neurotransmitters released from neurons bind to receptors expressed by astrocytes, activating a signaling pathway that leads to an increase in calcium concentration and the release of gliotransmitters that eventually modulate synaptic transmission. It is therefore likely that the activation of astrocytes impacts motor control. Here we review recent studies demonstrating that astrocytes inhibit, modulate, or trigger motor rhythmic behaviors.

DOI: 10.14814/phy2.15029 - 2021

PubReading [263] - Molecular mechanism of light-driven sodium pumping - K. Kovalev, V. Gordeliy et al.

Thu, 12 Jan 2023 13:00:22 GMT

The light-driven sodium-pumping rhodopsin KR2 from Krokinobacter eikastus is the only non- proton cation active transporter with demonstrated potential for optogenetics. However, the existing structural data on KR2 correspond exclusively to its ground state, and show no sodium inside the protein, which hampers the understanding of sodium-pumping mechanism. Here we present crystal structure of the O-intermediate of the physiologically relevant pentameric form of KR2 at the resolution of 2.1 Å, revealing a sodium ion near the retinal Schiff base, coordinated by N112 and D116 of the characteristic NDQ triad. We also obtained crystal structures of D116N and H30A variants, conducted metadynamics simulations and measured pumping activities of putative pathway mutants to demonstrate that sodium release likely proceeds alongside Q78 towards the structural sodium ion bound between KR2 protomers. Our findings highlight the importance of pentameric assembly for sodium pump function, and may be used for rational engineering of enhanced optogenetic tools.

https://doi.org/10.1038/s41467-020-16032-y - 2020

PubReading [262] - Lipids mediate supramolecular outer membrane protein assembly in bacteria - M. Webby, C. Kleanthous et al.

Wed, 11 Jan 2023 13:00:18 GMT

Beta Barrel outer membrane proteins (OMPs) cluster into supramolecular assemblies that give function to the outer membrane (OM) of Gram-negative bacteria. How such assemblies form is unknown. Here, through photoactivatable cross-linking into the Escherichia coli OM, coupled with simulations, and biochemical and biophysical analysis, we uncover the basis for OMP clustering in vivo. OMPs are typically surrounded by an annular shell of asymmetric lipids that mediate higher-order complexes with neighboring OMPs. OMP assemblies center on the abundant porins OmpF and OmpC, against which low-abundance monomeric Beta barrels, such as TonB-dependent transporters, are packed. Our study reveals OMP-lipid-OMP complexes to be the basic unit of supramolecular OMP assembly that, by extending across the entire cell surface, couples the requisite multifunctionality of the OM to its stability and impermeability.

doi: 10.1126/sciadv.adc9566 - 2022

PubReading [261] - Correlated Motions in Structural Biology - D. Xu, S. Meisburger, and N. Ando

Tue, 10 Jan 2023 13:00:36 GMT

Correlated motions in proteins arising from the collective movements of residues have long been proposed to be fundamentally important to key properties of proteins, from allostery and catalysis to evolvability. Recent breakthroughs in structural biology have made it possible to capture proteins undergoing complex conformational changes, yet intrinsic correlated motions within a conformation remain one of the least understood facets of protein structure. For many decades, the analysis of total X-ray scattering held the promise of animating crystal structures with correlated motions. With recent advances in both X-ray detectors and data interpretation methods, this long-held promise can now be met. In this Perspective, we will introduce how correlated motions are captured in total scattering and provide guidelines for the collection, interpretation, and validation of data. As structural biology continues to push the boundaries, we see an opportunity to gain atomistic insight into correlated motions using total scattering as a bridge between theory and experiment.

https://doi.org/10.1021/acs.biochem.1c00420 - 2021

PubReading [259] - Patient-Derived Organoids for Precision Cancer Immunotherapy - M. Groenholm, V. Cerullo et al.

Fri, 06 Jan 2023 13:00:38 GMT

Cancer immunotherapy has revolutionized the way tumors are treated. Nevertheless, efficient and robust testing platforms are still missing, including clinically relevant human ex vivo tumor assays that allow pretreatment testing of cancer therapies and selection of the most efficient and safe therapy for a specific patient. In the case of immunotherapy, this testing platform would require not only cancer cells, but also the tumor micro-environment, including immune cells. Here, we discuss the applications of patient-derived tumor organoid cultures and the possibilities in using complex immune–organoid cultures to provide preclinical testing platforms for precision cancer immunotherapy.

doi: 10.1158/0008-5472.CAN-20-4026 - 2021

PubReading [258] - Visualization of macromolecular structures - S. O’Donoghue, A. Olson et al.

Thu, 05 Jan 2023 12:00:59 GMT

Structural biology is rapidly accumulating a wealth of detailed information about protein function, binding sites, RNA, large assemblies and molecular motions. These data are increasingly of interest to a broader community of life scientists, not just structural experts. Visualization is a primary means for accessing and using these data, yet visualization is also a stumbling block that prevents many life scientists from benefiting from three-dimensional structural data. In this review, we focus on key biological questions where visualizing three-dimensional structures can provide insight and describe available methods and tools.

doi:10.1038/nmeth.1427 - 2010

PubReading [257] - AlphaFill- enriching AlphaFold models with ligands and cofactors - M. Hekkelman, A. Perrakis et al.

Wed, 04 Jan 2023 13:00:18 GMT

Artificial intelligence-based protein structure prediction approaches have had a transformative effect on biomolecular sciences. The predicted protein models in the AlphaFold protein structure database, however, all lack coordinates for small molecules, essential for molecular structure or function: hemoglobin lacks bound heme; zinc-finger motifs lack zinc ions essential for structural integrity and metalloproteases lack metal ions needed for catalysis. Ligands important for biological function are absent too; no ADP or ATP is bound to any of the ATPases or kinases. Here we present AlphaFill, an algorithm that uses sequence and structure similarity to ‘transplant’ such ‘missing’ small molecules and ions from experimentally determined structures to predicted protein models. The algorithm was successfully validated against experimental structures. A total of 12,029,789 transplants were performed on 995,411 AlphaFold models and are available together with associated validation metrics in the alphafill.eu databank, a resource to help scientists make new hypotheses and design targeted experiments.

https://doi.org/10.1038/s41592-022-01685-y - 2022

PubReading [256] - A Phenotypic Approach for the Identification of New Molecules for Targeted Protein Degradation Applications - P. Stacey, M. Muelbaier et al.

Tue, 03 Jan 2023 13:01:01 GMT

Targeted protein degradation is an emerging new strategy for the modulation of intracellular protein levels with applications in chemical biology and drug discovery. One approach to enable this strategy is to redirect the ubiquitin–proteasome system to mark and degrade target proteins of interest (POIs) through the use of proteolysis targeting chimeras (PROTACs). Although great progress has been made in enabling PROTACs as a platform, there are still a limited number of E3 ligases that have been employed for PROTAC design. Herein we report a novel phenotypic screening approach for the identification of E3 ligase binders. The key concept underlying this approach is the high-throughput modification of screening compounds with a chloroalkane moiety to generate HaloPROTACs in situ, which were then evaluated for their ability to degrade a GFP-HaloTag fusion protein in a cellular context. As proof of concept, we demonstrated that we could generate and detect functional HaloPROTACs in situ, using a validated Von Hippel–Lindau (VHL) binder that successfully degraded the GFP-HaloTag fusion protein in living cells. We then used this method to prepare and screen a library of approximately 2000 prospective E3 ligase-recruiting molecules.

DOI: 10.1177/2472555221017517 - 2021

PubReading [254] - DNA polymerase Epsilon and Delta exonuclease domain mutations in endometrial cancer - D. Church, I. Tomlinson

Fri, 30 Dec 2022 13:00:53 GMT

Accurate duplication of DNA prior to cell division is essential to suppress mutagenesis and tumour development. The high fidelity of eukaryotic DNA replication is due to a combination of accurate incorporation of nucleotides into the nascent DNA strand by DNA polymerases, the recognition and removal of mispaired nucleotides (proofreading) by the exonuclease activity of DNA polymerases Delta and Epsilon, and post-replication surveillance and repair of newly synthesized DNA by the mismatch repair (MMR) apparatus. While the contribution of defective MMR to neoplasia is well recognized, evidence that faulty DNA polymerase activity is important in cancer development has been limited. We have recently shown that germline POLE and POLD1 exonuclease domain mutations (EDMs) predispose to colorectal cancer (CRC) and, in the latter case, to endometrial cancer (EC). Somatic POLE mutations also occur in 5–10% of sporadic CRCs and underlie a hypermutator, microsatellite-stable molecular phenotype. We hypothesized that sporadic ECs might also acquire somatic POLE and/or POLD1 mutations. Here, we have found that missense POLE EDMs with good evidence of pathogenic effects are present in 7% of a set of 173 endometrial cancers, although POLD1 EDMs are uncommon. The POLE mutations localized to highly conserved residues and were strongly predicted to affect proofreading. Consistent with this, POLE-mutant tumours were hypermutated, with a high frequency of base substitutions, and an especially large relative excess of G:C>T:A transversions. All POLE EDM tumours were microsatellite stable, suggesting that defects in either DNA proofreading or MMR provide alternative mechanisms to achieve genomic instability and tumourigenesis.

doi:10.1093/hmg/ddt131 - 2013

PubReading [253] - Automated data collection for macromolecular crystallography - G. Winter & K. McAuley

Wed, 28 Dec 2022 19:06:36 GMT

An overview, together with some practical advice, is presented of the current status of the automation of macromolecular crystallography (MX) data collection, with a focus on MX beamlines at Diamond Light Source, UK.

doi:10.1016/j.ymeth.2011.06.010 - 2011

PubReading [252] - Epigenetic Heterogeneity in Friedreich Ataxia Underlies Variable FXN Reactivation - L. Rodden, S. Bidichandani et al.

Wed, 28 Dec 2022 12:00:13 GMT

Friedreich ataxia (FRDA) is typically caused by homozygosity for an expanded GAA triplet-repeat in intron 1 of the FXN gene. The expanded repeat induces repressive histone changes and DNA hypermethylation, which result in epigenetic silencing and FXN transcriptional deficiency. A class I histone deacetylase inhibitor (HDACi-109) reactivates the silenced FXN gene, although with considerable inter-individual variability, which remains etiologically unexplained. Because HDAC inhibitors work by reversing epigenetic silencing, we reasoned that epigenetic heterogeneity among patients may help to explain this inter-individual variability. As a surrogate measure for epigenetic heterogeneity, a highly quantitative measurement of DNA hypermethylation via bisulfite deep sequencing, with single molecule resolution, was used to assess the prevalence of unmethylated, partially methylated, and fully methylated somatic FXN molecules in PBMCs from a prospective cohort of 50 FRDA patients. Treatment of the same PBMCs from this cohort with HDACi-109 significantly increased FXN transcript to levels seen in asymptomatic heterozygous carriers, albeit with the expected inter-individual variability. Response to HDACi-109 correlated significantly with the prevalence of unmethylated and partially methylated FXN molecules, supporting the model that FXN reactivation involves a proportion of genes that are amenable to correction in non-dividing somatic cells, and that heavily methylated FXN molecules are relatively resistant to reactivation. FXN reactivation is a promising therapeutic strategy in FRDA, and inter-individual variability is explained, at least in part, by somatic epigenetic heterogeneity.

doi: 10.3389/fnins.2021.752921 - 2021

PubReading [251] - Directed DNA Metallization - G. Burley, T. Carell et al.

Tue, 27 Dec 2022 12:00:14 GMT

DNA is an outstanding material for the preparation of nano- and microscale assemblies, which are believed to have potential for the construction of nanoelectronic devices. DNA metallization procedures were developed in order to increase the conductivity of subsequent DNA nanostructures, thereby enabling their use as molecular wires.

10.1021/ja055517v - 2006

PubReading [249] - A flow platform for degradation-free CuAAC bioconjugation - M. Hatit, A. Watson et al.

Fri, 23 Dec 2022 12:00:28 GMT

The Cu-catalyzed azide-alkyne cycloaddition (CuAAC) reaction is a cornerstone method for the ligation of biomolecules. However, undesired Cu-mediated oxidation and Cu- contamination in bioconjugates limits biomedical utility. Here, we report a generic CuAAC flow platform for the rapid, robust, and broad-spectrum formation of discrete triazole bio-conjugates. This process leverages an engineering problem to chemical advantage: solvent-mediated Cu pipe erosion generates ppm levels of Cu in situ under laminar flow conditions. This is sufficient to catalyze the CuAAC reaction of small molecule alkynes and azides, fluorophores, marketed drug molecules, peptides, DNA, and therapeutic oligonucleotides. This flow approach, not replicated in batch, operates at ambient temperature and pressure, requires short residence times, avoids oxidation of sensitive functional groups, and produces products with very low ppm Cu contamination.

DOI: 10.1038/s41467-018-06551-0 - 2018

PubReading [248] - Third Generation Sequencing of Epigenetic DNA - B. Searle, M. Müller, T. Carell and A. Kellett

Thu, 22 Dec 2022 12:00:22 GMT

The discovery of epigenetic bases has revolutionised the understanding of disease and development. Among the most studied epigenetic marks are cytosines covalently modified at the 5 position. In order to gain insight into their biological significance, the ability to determine their spatiotemporal distribution within the genome is essential. Techniques for sequencing on ‘Next Generation’ platforms often involve harsh chemical treatments leading to sample degradation. Third generation sequencing promises to further revolutionise the field by providing long reads, enabling coverage of highly repetitive regions of the genome or structural variants considered unmappable by next generation sequencing technology. While the ability of third generation platforms to directly detect epigenetic modifications is continuously improving, at present chemical or enzymatic derivatisation presents the most convenient means of enhancing reliability. This review presents techniques available for the detection of cytosine modifications on third generation platforms.

https://doi.org/10.1002/anie.202215704 - 2022

PubReading [247] - The frequency of Ras mutations in cancer - I. Prior, F. Hood and J. Hartley

Wed, 21 Dec 2022 13:00:13 GMT

Ras is frequently mutated in cancer; however, there is a lack of consensus in the literature regarding the cancer mutation frequency of Ras, with quoted values varying from 10–30%. This variability is at least in part due to the selective aggregation of data from different databases and the dominant influence of particular cancer types and particular Ras isoforms within these datasets. In order to provide a more definitive figure for Ras mutation frequency in cancer, we cross-referenced the data in all major publicly accessible cancer mutation databases to determine reliable mutation frequency values for each Ras isoform in all major cancer types. These percentages were then applied to current US cancer incidence statistics to estimate the number of new patients each year that have Ras-mutant cancers. We find that ~19% of cancer patients harbor Ras mutations; equivalent to ~3.4 million new cases per year worldwide. We discuss the Ras isoform and mutation-specific trends evident within the datasets that are relevant to current Ras-targeted therapies.

doi:10.1158/0008-5472.CAN-19-3682 - 2020

PubReading [246] - Mechanistic insights into mRNA 3'-end processing - A. Kumar, M. Jinek et al.

Tue, 20 Dec 2022 13:00:13 GMT

The polyadenosine (poly(A)) tail found on the 3'-end of almost all eukaryotic mRNAs is important for mRNA stability and regulation of translation. mRNA 3'-end processing occurs co-transcriptionally and involves more than 20 proteins to specifically recognize the polyadenylation site, cleave the pre-mRNA, add a poly(A) tail, and trigger transcription termination. The polyadenylation site (PAS) defines the end of the 30-untranslated region (3'-UTR) and, therefore, selection of the cleavage site is a critical event in regulating gene expression. Integrated structural biology approaches including biochemical reconstitution of multi-subunit complexes, cross-linking mass spectrometry, and structural analyses by X- ray crystallography and single-particle electron cryo-microscopy (cryoEM) have enabled recent progress in understanding the molecular mechanisms of the mRNA 3'-end processing machinery. Here, we describe new molecular insights into pre-mRNA recognition, cleavage and polyadenylation.

https://doi.org/10.1016/j.sbi.2019.08.001 - 2019

PubReading [244] - The long and short of it - L. Passmore & T. Tang

Fri, 16 Dec 2022 12:00:13 GMT

Longer poly(A) tails improve translation in early development, but not in mature cells that have higher levels of the protein PABPC.

DOI: https://doi.org/10.7554/eLife.70757 - 2021

PubReading [243] - Liquid biopsies for residual disease and recurrence - J. Chee Ming Wan, N. Rosenfeld et al.

Thu, 15 Dec 2022 12:00:42 GMT

Detection of minimal residual disease in patients with cancer, who are in complete remission with no cancer cells detectable, has the potential to improve recurrence-free survival through treatment selection. Studies analyzing circulating tumor DNA (ctDNA) in patients with solid tumors suggest the potential to accurately predict and detect relapse, enabling treatment strategies that may improve clinical outcomes. Over the past decade, assays for ctDNA detection in plasma samples have steadily increased in sensitivity and specificity. These are applied for the detection of residual disease after treatment and for earlier detection of recurrence. Novel clinical trials are now assessing how assays for ‘‘residual disease and recurrence’’ (RDR) may influence current treatment paradigms and potentially change the landscape of risk classification for cancer recurrence. In this review, we appraise the progress of RDR detection using ctDNA and consider the emerging role of liquid biopsy in the monitoring and management of solid tumors.

https://doi.org/10.1016/j.medj.2021.11.001 - 2021

PubReading [242] - Single-particle cryo-EM at atomic resolution - T. Nakane, S. Scheres et al.

Wed, 14 Dec 2022 12:00:13 GMT

The three-dimensional positions of atoms in protein molecules define their structure and their roles in biological processes. The more precisely atomic coordinates are determined, the more chemical information can be derived and the more mechanistic insights into protein function may be inferred. Electron cryo-microscopy (cryo-EM) single-particle analysis has yielded protein structures with increasing levels of detail in recent years. However, it has proved difficult to obtain cryo-EM reconstructions with sufficient resolution to visualize individual atoms in proteins. Here we use a new electron source, energy filter and camera to obtain a 1.7 Å resolution cryo-EM reconstruction for a human membrane protein, the β3 GABAA receptor homopentamer. Such maps allow a detailed understanding of small-molecule coordination, visualization of solvent molecules and alternative conformations for multiple amino acids, and unambiguous building of ordered acidic side chains and glycans. Applied to mouse apoferritin, our strategy led to a 1.22 Å resolution reconstruction that offers a genuine atomic-resolution view of a protein molecule using single-particle cryo-EM. Moreover, the scattering potential from many hydrogen atoms can be visualized in difference maps, allowing a direct analysis of hydrogen-bonding networks. Our technological advances, combined with further approaches to accelerate data acquisition and improve sample quality, provide a route towards routine application of cryo-EM in high-throughput screening of small molecule modulators and structure-based drug discovery.

https://doi.org/10.1038/s41586-020-2829-0 - 2020

PubReading [241] - The neurons that restore walking after paralysis - C. Kathe, G. Courtine et al

Tue, 13 Dec 2022 12:00:13 GMT

A spinal cord injury interrupts pathways from the brain and brainstem that project to the lumbar spinal cord, leading to paralysis. Here we show that spatiotemporal epidural electrical stimulation (EES) of the lumbar spinal cord applied during neurorehabilitation (EESREHAB) restored walking in nine individuals with chronic spinal cord injury. This recovery involved a reduction in neuronal activity in the lumbar spinal cord of humans during walking. We hypothesized that this unexpected reduction reflects activity-dependent selection of specific neuronal subpopulations that become essential for a patient to walk after spinal cord injury. To identify these putative neurons, we modelled the technological and therapeutic features underlying EESREHAB in mice. We applied single-nucleus RNA sequencing and spatial transcriptomics to the spinal cords of these mice to chart a spatially resolved molecular atlas of recovery from paralysis. We then employed cell type and spatial prioritization to identify the neurons involved in the recovery of walking. A single population of excitatory interneurons nested within intermediate laminae emerged. Although these neurons are not required for walking before spinal cord injury, we demonstrate that they are essential for the recovery of walking with EES following spinal cord injury. Augmenting the activity of these neurons phenocopied the recovery of walking enabled by EESREHAB, whereas ablating them prevented the recovery of walking that occurs spontaneously after moderate spinal cord injury. We thus identified a recovery-organizing neuronal subpopulation that is necessary and sufficient to regain walking after paralysis. Moreover, our methodology establishes a framework for using molecular cartography to identify the neurons that produce complex behaviours.

https://doi.org/10.1038/s41586-022-05385-7 - 2022

PubReading [239] - Triplex-forming properties and enzymatic incorporation of a base-modified nucleotide capable of duplex DNA recognition at neutral pH - D. Rusling

Fri, 09 Dec 2022 12:00:23 GMT

The sequence-specific recognition of duplex DNA by unmodified parallel triplex-forming oligonucleotides is restricted to low pH conditions due to a necessity for cytosine protonation in the third strand. This has severely restricted their use as gene-targeting agents, as well as for the detection and/or functionalisation of synthetic or genomic DNA. Here I report that the nucleobase 6-amino-5-nitropyridin- 2-one (Z) finally overcomes this constraint by acting as an uncharged mimic of protonated cytosine. Synthetic TFOs containing the nucleobase enabled stable and selective triplex formation at oligopurine- oligopyrimidine sequences containing multiple isolated or contiguous GC base pairs at neutral pH and above. Moreover, I demonstrate a universal strategy for the enzymatic assembly of Z-containing TFOs using its commercially available deoxyribonucleotide triphosphate. These findings seek to improve not only the recognition properties of TFOs but also the cost and/or expertise associated with their chemical syntheses.

https://doi.org/10.1093/nar/gkab572 - 2021

PubReading [238] - Heart regeneration using pluripotent stem cells - S. Kadota, Y. Tanaka, Y. Shiba

Thu, 08 Dec 2022 12:00:13 GMT

Pluripotent stem cells (PSCs), which include embryonic and induced pluripotent stem cells (ESCs and iPSCs, respectively), have great potential in regenerative medicine for heart diseases due to their virtually unlimited cardiogenic capacity. Many preclinical studies have described the functional benefits after transplantation of PSC-derived cardiomyocytes (PSC-CMs). However, transient ventricular arrhythmias were detected after injection into non-human primates and swine ischemic hearts; as engrafted PSC-CMs form an electrical coupling between host and graft, the immature characteristics of PSC-CMs may serve as an ectopic pacemaker. We are entering a critical time in the development of novel therapies using PSC- CMs, with the recent first clinical trial using human iPSC-CMs (hiPSC-CMs) being launched in Japan. In this review, we summarize the updated knowledge, perspectives, and limitations of PSC-CMs for heart regeneration.

https://doi.org/10.1016/j.jjcc.2020.03.013 - 2020

PubReading [237] - Effect of the intratumoral microbiota on spatial and cellular heterogeneity in cancer - J. Niño, S. Bullman et al.

Wed, 07 Dec 2022 12:00:47 GMT

The tumour-associated microbiota is an intrinsic component of the tumour microenvironment across human cancer types1,2. Intratumoral host–microbiota studies have so far largely relied on bulk tissue analysis1–3, which obscures the spatial distribution and localized effect of the microbiota within tumours. Here, by applying in situ spatial-profiling technologies4 and single-cell RNA sequencing5 to oral squamous cell carcinoma and colorectal cancer, we reveal spatial, cellular and molecular host–microbe interactions. We adapted 10x Visium spatial transcriptomics to determine the identity and in situ location of intratumoral microbial communities within patient tissues. Using GeoMx digital spatial profiling6, we show that bacterial communities populate microniches that are less vascularized, highly immuno-suppressive and associated with malignant cells with lower levels of Ki-67 as compared to bacteria-negative tumour regions. We developed a single-cell RNA-sequencing method that we name INVADEseq (invasion–adhesion-directed expression sequencing) and, by applying this to patient tumours, identify cell-associated bacteria and the host cells with which they interact, as well as uncovering alterations in transcriptional pathways that are involved in inflammation, metastasis, cell dormancy and DNA repair. Through functional studies, we show that cancer cells that are infected with bacteria invade their surrounding environment as single cells and recruit myeloid cells to bacterial regions. Collectively, our data reveal that the distribution of the microbiota within a tumour is not random; instead, it is highly organized in microniches with immune and epithelial cell functions that promote cancer progression.

https://doi.org/10.1038/s41586-022-05435-0 - 2022

PubReading [236] - BioSimulators- a central registry of simulation engines and services for recommending specific tools - B. Shaikh, J. Karr et al.

Tue, 06 Dec 2022 13:00:23 GMT

Computational models have great potential to accelerate bioscience, bioengineering, and medicine. However, it remains challenging to reproduce and reuse simulations, in part, because the numerous formats and methods for simulating various subsystems and scales remain siloed by different software tools. For example, each tool must be executed through a distinct interface. To help investigators find and use simulation tools, we developed BioSimulators (https://biosimulators.org), a central registry of the capabilities of simulation tools and consistent Python, command-line and containerized interfaces to each version of each tool. The foundation of BioSimulators is standards, such as CellML, SBML, SED-ML and the COMBINE archive format, and validation tools for simulation projects and simulation tools that ensure these standards are used consistently. To help modelers find tools for particular projects, we have also used the registry to develop recommendation services. We anticipate that BioSimulators will help modelers exchange, reproduce, and combine simulations.

https://doi.org/10.1093/nar/gkac331 - 2022

PubReading [234] - The importance of synthetic chemistry in the pharmaceutical industry - K. Campos, E. Parmee et al.

Fri, 02 Dec 2022 12:00:45 GMT

Innovations in synthetic chemistry have enabled the discovery of many breakthrough therapies that have improved human health over the past century. In the face of increasing challenges in the pharmaceutical sector, continued innovation in chemistry is required to drive the discovery of the next wave of medicines. Novel synthetic methods not only unlock access to previously unattainable chemical matter, but also inspire new concepts as to how we design and build chemical matter. We identify some of the most important recent advances in synthetic chemistry as well as opportunities at the interface with partner disciplines that are poised to transform the practice of drug discovery and development.

DOI: 10.1126/science.aat0805 -2019

PubReading [233] - The importance of synthetic chemistry in the pharmaceutical industry - K. Campos, E. Parmee et al.

Thu, 01 Dec 2022 12:01:16 GMT

REVIEW SUMMARY

DOI: 10.1126/science.aat0805 -2019

PubReading [232] - The multiple flavors of GoU pairs in RNA - E. Westhof, M. Yusupov, G. Yusupova

Wed, 30 Nov 2022 12:00:18 GMT

Wobble GU pairs (or GoU) occur frequently within double‐stranded RNA helices interspersed within the standard G═C and A─U Watson‐Crick pairs. However, other types of GoU pairs interacting on their Watson‐Crick edges have been observed. The structural and functional roles of such alternative GoU pairs are surprisingly diverse and reflect the various pairings G and U can form by exploiting all the subtleties of their electronic configurations. Here, the structural characteristics of the GoU pairs are updated following the recent crystallographic structures of functional ribosomal complexes and the development in our understanding of ribosomal translation.

DOI: 10.1002/jmr.2782 - 2019

PubReading [231] - Light-activated communication in synthetic tissues - M. Booth, H. Bayley et al.

Tue, 29 Nov 2022 12:00:34 GMT

We have previously used three-dimensional (3D) printing to prepare tissue-like materials in which picoliter aqueous compartments are separated by lipid bilayers. These printed droplets are elaborated into synthetic cells by using a tightly regulated in vitro transcription/translation system. A light-activated DNA promoter has been developed that can be used to turn on the expression of any gene within the synthetic cells. We used light activation to express protein pores in 3D-printed patterns within synthetic tissues. The pores are incorporated into specific bilayer interfaces and thereby mediate rapid, directional electrical communication between subsets of cells. Accordingly, we have developed a functional mimic of neuronal transmission that can be controlled in a precise way.

DOI: 10.1126/sciadv.1600056 - 2016

PubReading [229] - RNA-Puzzles toolkit- a computational resource of RNA 3D structure benchmark datasets, structure manipulation, and evaluation tools - M. Magnus, Z. Miao et al.

Fri, 25 Nov 2022 12:00:19 GMT

Significant improvements have been made in the efficiency and accuracy of RNA 3D structure prediction methods during the succeeding challenges of RNA-Puzzles, a community-wide effort on the assessment of blind prediction of RNA tertiary structures. The RNA-Puzzles contest has shown, among others, that the development and validation of computational methods for RNA fold prediction strongly depend on the benchmark datasets and the structure comparison algorithms. Yet, there has been no systematic benchmark set or decoy structures available for the 3D structure prediction of RNA, hindering the standardization of comparative tests in the modelling of RNA structure. Furthermore, there has not been a unified set of tools that allows deep and complete RNA structure analysis, and at the same time, that is easy to use. Here, we present RNA-Puzzles toolkit, a computational resource including (i) decoy sets generated by different RNA 3D structure prediction methods (raw, for-evaluation and standardized datasets), (ii) 3D structure normalization, analysis, manipulation, visualization tools (RNA format, RNA normalizer, rna-tools) and (iii) 3D structure comparison metric tools (RNAQUA, MCQ4Structures). This resource provides a full list of computational tools as well as a standard RNA 3D structure prediction assessment protocol for the community.

doi: 10.1093/nar/gkz1108 - 2020

PubReading [228] - Data, data, burning deep, in the forests of the net - E. Westhof

Thu, 24 Nov 2022 12:00:28 GMT

Continuous and imaginative technological developments are leading to a massive accumulation of various types of data in all areas of biological research. As a result, the central importance of databases is increasing. Databases related to biology must not only be structured using controlled vocabularies, but also be fully integrated into the whole biological domain. To achieve this goal, they must be systematically grounded in biological evolution and exploit the available tools of evolutionary systematics to contribute to our understanding of life processes.

https://doi.org/10.1016/j.bbrc.2022.09.030 - 2022

PubReading [227] - Artificial intelligence in chemistry and drug design - N. Brown, N. Schneider et al

Wed, 23 Nov 2022 12:00:37 GMT

The discovery of molecular structures with desired properties for applications in drug discovery, crop protection, or chemical biology is among the most impactful scientific challenges. However, given the complexity of biological systems and the associated cost for experiments and trials, molecular design is also scientifically very challenging, prone to failure, inherently expensive and time consuming. To improve our odds and the timelines in this process, and to identify good starting points, unbiased incorporation of knowledge through continuous analysis of literature and patents from different scientific fields is required

https://doi.org/10.1007/s10822-020-00317-x - 2020

PubReading [226] - The threat of programmed DNA damage to neuronal genome integrity and plasticity - K. Caldecott, M. Ward and A. Nussenzweig

Tue, 22 Nov 2022 12:01:17 GMT

The neuronal genome is particularly sensitive to loss or attenuation of DNA repair, and many neurological diseases ensue when DNA repair is impaired. It is well-established that the neuronal genome is subjected to stochastic DNA damage, most likely because of extensive oxidative stress in the brain. However, recent studies have identified unexpected high levels of ‘programmed’ DNA breakage in neurons, which we propose arise during physiological DNA metabolic processes intrinsic to neuronal development, differentiation and maintenance. The role of programmed DNA breaks in normal neuronal physiology and disease remains relatively unexplored thus far. However, bulk and single-cell sequencing analyses of neurodegenerative diseases have revealed age-related somatic mutational signatures that are enriched in regulatory regions of the genome. Here, we explore a paradigm of DNA repair in neurons, in which the genome is safeguarded from erroneous impacts of programmed genome breakage intrinsic to normal neuronal function.

https://doi.org/10.1038/s41588-021-01001-y - 2022

PubReading [224] - Drug Design- Where We Are and Future Prospects - G. Zagotto and M. Bortoli

Fri, 18 Nov 2022 12:00:29 GMT

Medicinal chemistry is facing new challenges in approaching precision medicine. Several powerful new tools or improvements of already used tools are now available to medicinal chemists to help in the process of drug discovery, from a hit molecule to a clinically used drug. Among the new tools, the possibility of considering folding intermediates or the catalytic process of a protein as a target for discovering new hits has emerged. In addition, machine learning is a new valuable approach helping medicinal chemists to discover new hits. Other abilities, ranging from the better understanding of the time evolution of biochemical processes to the comprehension of the biological meaning of the data originated from genetic analyses, are on their way to progress further in the drug discovery field toward improved patient care. In this sense, the new approaches to the delivery of drugs targeted to the central nervous system, together with the advancements in understanding the metabolic pathways for a growing number of drugs and relating them to the genetic characteristics of patients, constitute important progress in the field.

https://doi.org/10.3390/molecules26227061 - 2021

PubReading [223] - The Discovery of the DNA Double Helix - A. Klung

Thu, 17 Nov 2022 12:01:00 GMT

Fifty years ago, on 25th April 1953, there appeared three papers in the journal, Nature, which changed our view of the world. The structure of the DNA double helix, with its complementary base-pairing, is one of the greatest discoveries in biology in the 20th Century. It was also most dramatic, since, quite unexpectedly, the structure itself pointed to the way in which a DNA molecule might replicate itself, and hence revealed the “secret of life”. The structure was solved in the Cavendish Laboratory, Cambridge by Francis Crick and James Watson, using X-ray diffraction data from fibres of DNA obtained by Rosalind Franklin at King’s College, London.

doi:10.1016/j.jmb.2003.11.015 - 2004

PubReading [222] - Click Chemistry Enables Rapid Amplification of Full-Length Reverse Transcripts for Long-Read Third Generation Sequencing - E. Schönegger, T. Frischmuth

Wed, 16 Nov 2022 13:00:48 GMT

Here we describe the development of a novel click chemistry-based method for the generation and amplification of full-length cDNA libraries from total RNA, while avoiding the need for problematic template-switching (TS) reactions. Compared with prior efforts, our method involves neither random priming nor stochastic cDNA termination, thus enabling amplification of transcripts that were previously inaccessible via related click chemistry-based RNA sequencing techniques. A key modification involving the use of PCR primers containing two overhanging 3'-nucleotides substantially improved the read-through compatibility of the 1,4-disubstituted 1,2,3-triazole-containing cDNA, where such modifications typically hinder amplification. This allowed us to more than double the possible insert size compared with the state-of-the art click chemistry-based technique, PAC-seq. Furthermore, our method performed on par with a commercially available PCR-cDNA RNA sequencing kit, as determined by Oxford Nanopore sequencing. Given the known advantages of PAC-seq, namely, suppression of PCR artifacts, we anticipate that our contribution could enable diverse applications including improved analyses of mRNA splicing variants and fusion transcripts.

DOI: 10.1021/acs.bioconjchem.2c00353 - 2022

PubReading [221] - Hairpin-like siRNA-Based Spherical Nucleic Acids - M. Vasher, G. Yamankurt, C. Mirkin

Tue, 15 Nov 2022 12:00:29 GMT

The therapeutic use of small interfering RNAs (siRNAs) as gene regulation agents has been limited by their poor stability and delivery. Although arranging siRNAs into a spherical nucleic acid (SNA) architecture to form siRNA-SNAs increases their stability and uptake, prototypical siRNA-SNAs consist of a hybridized architecture that causes guide strand dissociation from passenger strands, which limits the delivery of active siRNA duplexes. In this study, a new SNA design that directly attaches both siRNA strands to the SNA core through a single hairpin-shaped molecule to prevent guide strand dissociation is introduced and investigated. This hairpin-like architecture increases the number of siRNA duplexes that can be loaded onto an SNA by 4-fold compared to the original hybridized siRNA-SNA architecture. As a result, the hairpin-like siRNA-SNAs exhibit a 6-fold longer half-life in serum and decreased cytotoxicity. In addition, the hairpin-like siRNA-SNA produces more durable gene knockdown than the hybridized siRNA-SNA. This study shows how the chemistry used to immobilize siRNA on nanoparticles can markedly enhance biological function, and it establishes the hairpin-like architecture as a next-generation SNA construct that will be useful in life science and medical research.

DOI: 10.1021/jacs.1c12750 - 2022

PubReading [220] - Interbase FRET in RNA: from A to Z - A. Fuechtbauer, M. Wilhelmsson et al.

Mon, 14 Nov 2022 12:00:19 GMT

Interbase FRET can reveal highly detailed information about distance, orientation and dynamics in nucleic acids, complementing the existing structure and dynamics techniques. We here report the first RNA base analogue FRET pair, consisting of the donor tCO and the non-emissive acceptor tCnitro. The acceptor ribonucleoside is here synthesised and incorporated into RNA for the first time. This FRET pair accurately reports the average structure of A-form RNA, and its utility for probing RNA structural changes is demonstrated by monitoring the transition from A- to Z-form RNA. Finally, the measured FRET data were compared with theoretical FRET patterns obtained from two previously reported Z-RNA PDB structures, to shed new light on this elusive RNA conformation.

doi: 10.1093/nar/gkz812 - 2019

PubReading [219] - An integrated workflow for crosslinking mass spectrometry - M. Mendes, J. Rappsilber et al

Fri, 11 Nov 2022 13:00:23 GMT

We present a concise workflow to enhance the mass spectrometric detection of crosslinked peptides by introducing sequential digestion and the crosslink identification software xiSEARCH. Sequential digestion enhances peptide detection by selective shortening of long tryptic peptides. We demonstrate our simple 12-fraction protocol for crosslinked multi-protein complexes and cell lysates, quantitative analysis, and high-density crosslinking, without requiring specific crosslinker features. This overall approach reveals dynamic protein-protein interaction sites, which are accessible, have fundamental functional relevance and are therefore ideally suited for the development of small molecule inhibitors.

DOI: 10.15252/msb.20198994 - 2019

PubReading [218] - Covalent labeling of immune cells - L. Rösner, C. Konken, D. Depke, A. Rentmeister, M. Schäfers

Thu, 10 Nov 2022 12:00:27 GMT

Inflammation is a common, fast, and innate response of the immune system to sterile or infectious tissue damage or autoimmune triggers. It aims at minimizing tissue destruction and maintaining organ function, hence is vital to life. Therefore, the immune system comprises the concerted action of a variety of different immune cells with specific tasks in the initiation, maintenance, and termination of inflammation. Visualizing their localization, trafficking, and interaction is of utmost importance to unravel the dynamics of inflammation in the living organism and requires tools for cell-specific labeling and imaging. Many concepts for covalent cell-type or protein-specific labeling have been developed, but only few have been implemented for labeling immune cells. Here, we review approaches that were already successful for fluorescent reporters and radioactive nuclides. We also provide a glimpse on emerging technologies that bear potential for immune cell labeling and imaging in vivo.

doi: 10.1126/scitranslmed.3007094 - 2014

PubReading [217] - Detection of Circulating Tumor DNA in Early- and Late-Stage Human Malignancies - C. Bettegowda, L. Diaz Jr et al

Wed, 09 Nov 2022 12:00:30 GMT

The development of noninvasive methods to detect and monitor tumors continues to be a major challenge in oncology. We used digital polymerase chain reaction-based technologies to evaluate the ability of circulating tumor DNA (ctDNA) to detect tumors in 640 patients with various cancer types. We found that ctDNA was detectable in >75% of patients with advanced pancreatic, ovarian, colorectal, bladder, gastroesophageal, breast, melanoma, hepatocellular, and head and neck cancers, but in less than 50% of primary brain, renal, prostate, or thyroid cancers. In patients with localized tumors, ctDNA was detected in 73, 57, 48, and 50% of patients with colorectal cancer, gastroesophageal cancer, pancreatic cancer, and breast adenocarcinoma, respectively. ctDNA was often present in patients without detectable circulating tumor cells, suggesting that these two biomarkers are distinct entities. In a separate panel of 206 patients with metastatic colorectal cancers, we showed that the sensitivity of ctDNA for detection of clinically relevant KRAS gene mutations was 87.2% and its specificity was 99.2%. Finally, we assessed whether ctDNA could provide clues into the mechanisms underlying resistance to epidermal growth factor receptor blockade in 24 patients who objectively responded to therapy but subsequently relapsed. Twenty-three (96%) of these patients developed one or more mutations in genes involved in the mitogen-activated protein kinase pathway. Together, these data suggest that ctDNA is a broadly applicable, sensitive, and specific biomarker that can be used for a variety of clinical and research purposes in patients with multiple different types of cancer.

doi: 10.1126/scitranslmed.3007094. - 2014

PubReading [216] - Bottom-Up Assembly of Synthetic Cells with a DNA Cytoskeleton - K. Jahnke, K. Göpfrich et al

Tue, 08 Nov 2022 12:00:44 GMT

Cytoskeletal elements, like actin and myosin, have been reconstituted inside lipid vesicles toward the vision to reconstruct cells from the bottom up. Here, we realize the de novo assembly of entirely artificial DNA-based cytoskeletons with programmed multifunctionality inside synthetic cells. Giant unilamellar lipid vesicles (GUVs) serve as cell-like compartments, in which the DNA cytoskeletons are repeatedly and reversibly assembled and disassembled with light using the cis–trans isomerization of an azobenzene moiety positioned in the DNA tiles. Importantly, we induced ordered bundling of hundreds of DNA filaments into more rigid structures with molecular crowders. We quantify and tune the persistence length of the bundled filaments to achieve the formation of ring-like cortical structures inside GUVs, resembling actin rings that form during cell division. Additionally, we show that DNA filaments can be programmably linked to the compartment periphery using cholesterol-tagged DNA as a linker. The linker concentration determines the degree of the cortex-like network formation, and we demonstrate that the DNA cortex-like network can deform GUVs from within. All in all, this showcases the potential of DNA nanotechnology to mimic the diverse functions of a cytoskeleton in synthetic cells.

https://doi.org/10.1021/acsnano.1c10703 - 2022

PubReading [115] - Therapeutic application of sequence-specific binding molecules for novel genome editing tools - J. Nakao, T. Yamamoto and A. Yamayoshi

Mon, 07 Nov 2022 12:01:11 GMT

Genome editing has been expected to widely increase the available treatment options for various diseases and permit pharmaceutical interventions in previously untreatable conditions. The availability of genome editing tools was dramatically increased by the development of the CRISPR-Cas9 system. However, a number of issues limit the use of the CRISPR-Cas9 system and other gene-editing tools in the clinical treatment of diseases. This review summarized the history and types of genome editing tools and limitations of their use. In addition, the study addressed several next-generation technologies aiming to overcome the limitations of current gene therapy protocols in an effort to accelerate the clinical development of potential treatment options. This review has provided an extensive foundation of the current state of genome editing technology and its clinical development. This review also indicate that the study additionally highlighted the need for multidisciplinary approaches to overcome current bottlenecks in the development of genome editing.

https://doi.org/10.1016/j.dmpk.2021.100427 - 2022

PubReading [214] - Parallel CRISPR-Cas9 screens clarify impacts of p53 on screen performance - A. Bowden, S. Jackson et al.

Fri, 04 Nov 2022 21:00:19 GMT

CRISPR-Cas9 genome engineering has revolutionised high-throughput functional genomic screens. However, recent work has raised concerns regarding the performance of CRISPR- Cas9 screens using TP53 wild-type human cells due to a p53-mediated DNA damage response (DDR) limiting the efficiency of generating viable edited cells. To directly assess the impact of cellular p53 status on CRISPR-Cas9 screen performance, we carried out parallel CRISPR-Cas9 screens in wild-type and TP53 knockout human retinal pigment epithelial cells using a focused dual guide RNA library targeting 852 DDR-associated genes. Our work demonstrates that although functional p53 status negatively affects identification of significantly depleted genes, optimal screen design can nevertheless enable robust screen performance. Through analysis of our own and published screen data, we highlight key factors for successful screens in both wild-type and p53-deficient cells.

DOI: https://doi.org/10.7554/eLife.55325 - 2020

PubReading [213] - Stabilisation of self-assembled DNA crystals by triplex-directed photo-cross-linking - H. Abdallah, N. Seeman et al.

Thu, 03 Nov 2022 21:00:18 GMT

The tensegrity triangle is a robust DNA motif that can self-assemble to generate macroscopic three-dimensional crystals. However, the stability of these crystals is dependent on the high ionic conditions used for crystal growth. Here we demonstrate that a triplex-forming oligonucleotide can be used to direct the specific intercalation, and subsequent photo-cross-linking, of 4,5′,8-trimethylpsoralen to single or multiple loci within or between the tiles of the crystal. Cross-linking between the tiles of the crystal improves their thermal stability. Such an approach is likely to facilitate the removal of crystals from their mother liquor and may prove useful for applications that require greater crystal stability.

DOI: 10.1039/c6cc03695c - 2016

PubReading [212] - Glyoxal oxidases: their nature and properties - M. Daou and C.Faulds

Wed, 02 Nov 2022 21:00:18 GMT

H2O2 has been found to be required for the activity of the main microbial enzymes responsible for lignin oxidative cleavage, peroxidases. Along with other small radicals, it is implicated in the early attack of plant biomass by fungi. Among the few extracellular H2O2-generating enzymes known are the glyoxal oxidases (GLOX). GLOX is a copper-containing enzyme, sharing high similarity at the level of active site structure and chemistry with galactose oxidase. Genes encoding GLOX enzymes are widely distributed among wood-degrading fungi especially white-rot degraders, plant pathogenic and symbiotic fungi. GLOX has also been identified in plants. Although widely distributed, only few examples of characterized GLOX exist. The first characterized fungal GLOX was isolated from Phanerochaete chrysosporium. The GLOX from Utilago maydis has a role in filamentous growth and pathogenicity. More recently, two other glyoxal oxidases from the fungus Pycnoporus cinnabarinus were also characterized. In plants, GLOX from Vitis pseudoreticulata was found to be implicated in grapevine defence mechanisms. Fungal GLOX were found to be activated by peroxidases in vitro suggesting a synergistic and regulatory relationship between these enzymes. The substrates oxidized by GLOX are mainly aldehydes generated during lignin and carbohydrates degradation. The reactions catalysed by this enzyme such as the oxidation of toxic molecules and the production of valuable compounds (organic acids) makes GLOX a promising target for biotechnological applications. This aspect on GLOX remains new and needs to be investigated.

DOI 10.1007/s11274-017-2254-1 - 2017

PubReading [211] - 3D DNA Crystals and Nanotechnology - P. Paukstelis and N. Seeman

Tue, 01 Nov 2022 12:00:45 GMT

DNA’s molecular recognition properties have made it one of the most widely used biomacromolecular construction materials. The programmed assembly of DNA oligonucleotides has been used to create complex 2D and 3D self-assembled architectures and to guide the assembly of other molecules. The origins of DNA nanotechnology are rooted in the goal of assembling DNA molecules into designed periodic arrays, i.e., crystals. Here, we highlight several DNA crystal structures, the progress made in designing DNA crystals, and look at the current prospects and future directions of DNA crystals in nanotechnology.

doi:10.3390/cryst6080097 - 2016

PubReading [210] - Interbase‐FRET binding assay for pre‐microRNAs - M. Bood, M. Wilhelmsson, M. Grøtli et al.

Mon, 31 Oct 2022 12:00:19 GMT

The aberrant expression of microRNAs (miRs) has been linked to several human diseases. A promising approach for targeting these anomalies is the use of small‐molecule inhibitors of miR biogenesis. These inhibitors have the potential to (i) dissect miR mechanisms of action, (ii) discover new drug targets, and (iii) function as new therapeutic agents. Here, we designed Förster resonance energy transfer (FRET)‐labeled oligoribonucleotides of the precursor of the oncogenic miR‐21 (pre‐miR‐21) and used them together with a set of aminoglycosides to develop an interbase‐FRET assay to detect ligand binding to pre‐miRs. Our interbase‐FRET assay accurately reports structural changes of the RNA oligonucleotide induced by ligand binding. We demonstrate its application in a rapid, qualitative drug candidate screen by assessing the relative binding affinity between 12 aminoglycoside antibiotics and pre‐miR‐21. Surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) were used to validate our new FRET method, and the accuracy of our FRET assay was shown to be similar to the established techniques. With its advantages over SPR and ITC owing to its high sensitivity, small sample size, straightforward technique and the possibility for high‐throughput expansion, we envision that our solution‐based method can be applied in pre‐miRNA–target binding studies.

https://doi.org/10.1038/s41598-021-88922-0 - 2021

PubReading [209] - Flipping the Switch: Innovations in Inducible Probes for Protein Profiling - S. McKenna, E. Fay and J. McGouran

Fri, 28 Oct 2022 11:00:17 GMT

Over the past two decades, activity-based probes have enabled a range of discoveries, including the characterization of new enzymes and drug targets. However, their suitability in some labeling experiments can be limited by nonspecific reactivity, poor membrane permeability, or high toxicity. One method for overcoming these issues is through the development of “inducible” activity-based probes. These probes are added to samples in an unreactive state and require in situ transformation to their active form before labeling can occur. In this Review, we discuss a variety of approaches to inducible activity-based probe design, different means of probe activation, and the advancements that have resulted from these applications. Additionally, we highlight recent developments which may provide opportunities for future inducible activity-based probe innovations.

https://doi.org/10.1021/acschembio.1c00572 - 2021

PubReading [208] - Cystatin C as a biomarker of chronic kidney disease: latest developments - S. Benoit, E. Ciccia, P. Devarajan

Thu, 27 Oct 2022 11:00:17 GMT

Chronic kidney disease (CKD) is common, occurring in over 10% of individuals globally, and is increasing in prevalence. The limitations of traditional biomarkers of renal dysfunction, such as serum creatinine, have been well-demonstrated in the literature. Therefore, augmenting clinical assessment with newer biomarkers, such as serum cystatin C, has the potential to improve disease monitoring and patient care. The present paper assesses the utility and limitations of serum cystatin C as a biomarker for CKD in light of the current literature. Serum cystatin C has been well established as an early and accurate biomarker of CKD that is particularly helpful in patients for whom creatinine is an inadequate marker or for whom more cumbersome methods of glomerular filtration rate (GFR) measurement are impractical. Current research questions are no longer focused on if, but rather when and how often cystatin C should be used in the evaluation of CKD patients. However, transition of all reagents and estimated GFR equations to the newly established International Standard is critical for developing generalizable data.

doi:10.1080/14737159.2020.1768849 - 2020

PubReading [207] - Holliday junction-resolving enzymes—structures and mechanisms - David Lilley

Wed, 26 Oct 2022 11:00:18 GMT

Holliday junction-resolving enzymes are nucleases that are highly specific for the structure of the junction, to which they bind in dimeric form. Two symmetrically disposed cleavages are made. These are not simultaneous, but the second cleavage is accelerated relative to the first, so ensuring that bilateral cleavage occurs during the lifetime of the DNA–protein complex. In eukaryotic cells there are two known junction-resolving activities. GEN1 is similar to enzymes from lower organisms. A crystallographic structure of a fungal GEN1 bound to the product of resolution has been determined. These complexes are dimerized within the crystal lattice such that the strands of the products may be simply reconnected to form a junction. These structures suggest a trajectory for the resolution process.

doi:10.1002/1873-3468.12529 - 2016

PubReading [206] - The path for metal complexes to a DNA target - A. Komora and J. Barton

Tue, 25 Oct 2022 11:00:38 GMT

The discovery of cisplatin as a therapeutic agent stimulated a new era in the application of transition metal complexes for therapeutic design. Here we describe recent results on a variety of transition metal complexes targeted to DNA to illustrate many of the issues involved in new therapeutic design. We describe first structural studies of complexes bound covalently and non-covalently to DNA to identify potential lesions within the cell. We then review the biological fates of these complexes, illustrating the key elements in obtaining potent activity, the importance of uptake and subcellular localization of the complexes, as well as the techniques used to delineate these characteristics. Genomic DNA provides a challenging but valuable target for new transition metal-based therapeutics.

doi:10.1039/c3cc00177f - 2013

PubReading [205] - Targeting tumor cell senescence and polyploidy as potential therapeutic strategies - T. Saleh, D. Gewirtz et al.

Mon, 24 Oct 2022 11:00:19 GMT

Senescence is a unique state of growth arrest that develops in response to a plethora of cellular stresses, including replicative exhaustion, oxidative injury, and genotoxic insults. Senescence has been implicated in the pathogenesis of multiple aging-related pathologies, including cancer. In cancer, senescence plays a dual role, initially acting as a barrier against tumor progression by enforcing a durable growth arrest in premalignant cells, but potentially promoting malignant transformation in neighboring cells through the secretion of pro-tumorigenic drivers. Moreover, senescence is induced in tumor cells upon exposure to a wide variety of conventional and targeted anticancer drugs (termed Therapy-Induced Senescence—TIS), representing a critical contributing factor to therapeutic outcomes. As with replicative or oxidative senescence, TIS manifests as a complex phenotype of macromolecular damage, energetic dysregulation, and altered gene expression. Senescent cells are also frequently polyploid. In vitro studies have suggested that polyploidy may confer upon senescent tumor cells the ability to escape from growth arrest, thereby providing an additional avenue whereby tumor cells escape the lethality of anticancer treatment. Polyploidy in tumor cells is also associated with persistent energy production, chromatin remodeling, self-renewal, stemness and drug resistance - features that are also associated with escape from senescence and conversion to a more malignant phenotype. However, senescent cells are highly heterogenous and can present with variable phenotypes, where polyploidy is one component of a complex reversion process. Lastly, emerging efforts to pharmacologically target polyploid tumor cells might pave the way towards the identification of novel targets for the elimination of senescent tumor cells by the incorporation of senolytic agents into cancer therapeutic strategies.

https://doi.org/10.1016/j.semcancer.2020.12.010 -2020

PubReading [204] - Antibody targeting of E3 ubiquitin ligases for receptor degradation - H. Marei, F. de Sousa e Melo et al.

Fri, 21 Oct 2022 11:00:26 GMT

Most current therapies that target plasma membrane receptors function by antagonizing ligand binding or enzymatic activities. However, typical mammalian proteins comprise multiple domains that execute discrete but coordinated activities. Thus, inhibition of one domain often incompletely suppresses the function of a protein. Indeed, targeted protein degradation technologies, including proteolysis-targeting chimeras1 (PROTACs), have highlighted clinically important advantages of target degradation over inhibition2. However, the generation of heterobifunctional compounds binding to two targets with high affinity is complex, particularly when oral bioavailability is required3. Here we describe the development of proteolysis-targeting antibodies (PROTABs) that tether cell-surface E3 ubiquitin ligases to transmembrane proteins, resulting in target degradation both in vitro and in vivo. Focusing on zinc- and ring finger 3 (ZNRF3), a Wnt-responsive ligase, we show that this approach can enable colorectal cancer-specific degradation. Notably, by examining a matrix of additional cell-surface E3 ubiquitin ligases and transmembrane receptors, we demonstrate that this technology is amendable for ‘on-demand’ degradation. Furthermore, we offer insights on the ground rules governing target degradation by engineering optimized antibody formats. In summary, this work describes a strategy for the rapid development of potent, bioavailable and tissue-selective degraders of cell-surface proteins.

https://doi.org/10.1038/s41586-022-05235-6 - 2022

PubReading [203] - The mechanism of replication stalling and recovery within repetitive DNA - C. Cases-Delucchi, M. Daza-Martin, S. Williams & G. Coster

Thu, 20 Oct 2022 11:00:18 GMT

Accurate chromosomal DNA replication is essential to maintain genomic stability. Genetic evidence suggests that certain repetitive sequences impair replication, yet the underlying mechanism is poorly defined. Replication could be directly inhibited by the DNA template or indirectly, for example by DNA-bound proteins. Here, we reconstitute replication of mono-, di- and trinucleotide repeats in vitro using eukaryotic replisomes assembled from purified proteins. We find that structure-prone repeats are sufficient to impair replication. Whilst template unwinding is unaffected, leading strand synthesis is inhibited, leading to fork uncoupling. Synthesis through hairpin-forming repeats is rescued by replisome-intrinsic mechanisms, whereas synthesis of quadruplex-forming repeats requires an extrinsic accessory helicase. DNA-induced fork stalling is mechanistically similar to that induced by leading strand DNA lesions, highlighting structure-prone repeats as an important potential source of replication stress. Thus, we propose that our understanding of the cellular response to replication stress may also be applied to DNA-induced replication stalling.

https://doi.org/10.1038/s41467-022-31657-x - 2022

PubReading [202] - Potentialities of Gene Therapy in Pediatric Endocrinology - G. Frontino. M. Stancampiano, A. Aiuti

Wed, 19 Oct 2022 11:01:00 GMT

Gene therapy has become an appealing therapeutic option in many pediatric fields, including endocrinology. Unlike traditional drugs based on molecules that require repeated and frequent burdensome administrations, a single genetic therapeutic intervention may allow durable and curative clinical benefits. Although this highly innovative technology holds a great promise for the treatment of monogenic diseases, its clinical applications in the field of endocrinology have been so far challenging. In this review, we will discuss various ex vivo and in vivo approaches and potential applications of gene addition and gene editing approaches for treating hyperfunctional and hypofunctional endocrine diseases due to intrinsic defects or autoimmune origin. We will focus on the recent advances in gene therapy approaches aimed at treating type 1 diabetes and monogenic forms of endocrinopa- thies such as growth hormone deficiency, congenital adrenal hyperplasia, diabetes insipidus, IPEX, as well as their trends and future direction.

DOI: 10.1159/000520965 - 2021

PubReading [201] - The cryo-EM method microcrystal electron diffraction (MicroED) - B. Nannenga and T. Gone

Tue, 18 Oct 2022 20:00:17 GMT

In 2013 we established a cryo-electron microscopy (cryo-EM) technique called microcrystal electron diffraction (MicroED). Since that time, data collection and analysis schemes have been fine-tuned, and structures for more than 40 different proteins, oligopeptides and organic molecules have been determined. Here we review the MicroED technique and place it in context with other structure-determination methods. We showcase example structures solved by MicroED and provide practical advice to prospective users.

doi.org/10.1038/s41592-019-0395-x - 2019

PubReading [200] - Stealth Fluorescence Labeling for Live Microscopy Imaging of mRNA Delivery - T. Baladi, M. Wilhelmsson et al

Mon, 17 Oct 2022 11:00:49 GMT

Methods for tracking RNA inside living cells without perturbing their natural interactions and functions are critical within biology and, in particular, to facilitate studies of therapeutic RNA delivery. We present a stealth labeling approach that can efficiently, and with high fidelity, generate RNA transcripts, through enzymatic incorporation of the triphosphate of tCO, a fluorescent tricyclic cytosine analogue. We demonstrate this by incorporation of tCO in up to 100% of the natural cytosine positions of a 1.2 kb mRNA encoding for the histone H2B fused to GFP (H2B:GFP). Spectroscopic characterization of this mRNA shows that the incorporation rate of tCO is similar to cytosine, which allows for efficient labeling and controlled tuning of labeling ratios for different applications. Using live cell confocal microscopy and flow cytometry, we show that the tCO-labeled mRNA is efficiently translated into H2B:GFP inside human cells. Hence, we not only develop the use of fluorescent base analogue labeling of nucleic acids in live-cell microscopy but also, importantly, show that the resulting transcript is translated into the correct protein. Moreover, the spectral properties of our transcripts and their translation product allow for their straightforward, simultaneous visualization in live cells. Finally, we find that chemically transfected tCO-labeled RNA, unlike a state- of-the-art fluorescently labeled RNA, gives rise to expression of a similar amount of protein as its natural counterpart, hence representing a methodology for studying natural, unperturbed processing of mRNA used in RNA therapeutics and in vaccines, like the ones developed against SARS-CoV-2.

https://doi.org/10.1021/jacs.1c00014 - 2021

PubReading [199] - Live-seq enables temporal transcriptomic recording of single cells - W. Chen, B. Deplanke et al

Fri, 14 Oct 2022 11:00:17 GMT

Single-cell transcriptomics (scRNA-seq) has greatly advanced our ability to characterize cellular heterogeneity1. However, scRNA-seq requires lysing cells, which impedes further molecular or functional analyses on the same cells. Here, we established Live-seq, a single-cell transcriptome profiling approach that preserves cell viability during RNA extraction using fluidic force microscopy2,3, thus allowing to couple a cell’s ground-state transcriptome to its downstream molecular or phenotypic behaviour. To benchmark Live-seq, we used cell growth, functional responses and whole-cell transcriptome read-outs to demonstrate that Live-seq can accurately stratify diverse cell types and states without inducing major cellular perturbations. As a proof of concept, we show that Live-seq can be used to directly map a cell’s trajectory by sequentially profiling the transcriptomes of individual macrophages before and after lipopolysaccharide (LPS) stimulation, and of adipose stromal cells pre- and post-differentiation. In addition, we demonstrate that Live-seq can function as a transcriptomic recorder by preregistering the transcriptomes of individual macrophages that were subsequently monitored by time-lapse imaging after LPS exposure. This enabled the unsupervised, genome-wide ranking of genes on the basis of their ability to affect macrophage LPS response heterogeneity, revealing basal Nfkbia expression level and cell cycle state as important phenotypic determinants, which we experimentally validated. Thus, Live-seq can address a broad range of biological questions by transforming scRNA-seq from an end-point to a temporal analysis approach.

https://doi.org/10.1038/s41586-022-05046-9 - 2022

PubReading [198] - Neutron crystallography: opportunities, challenges, and limitations - M. Blakeley, A. Podjarny et al

Thu, 13 Oct 2022 11:00:27 GMT

Neutron crystallography has had an important, but relatively small role in structural biology over the years. In this review of recently determined neutron structures, a theme emerges of a field currently expanding beyond its traditional boundaries, to address larger and more complex problems, with smaller samples and shorter data collection times, and employing more sophisticated structure determination and refinement methods. The origin of this transformation can be found in a number of advances including first, the development of neutron image-plates and quasi-Laue methods at nuclear reactor neutron sources and the development of time-of-flight Laue methods and electronic detectors at spallation neutron sources; second, new facilities and methods for sample perdeuteration and crystallization; third, new approaches and computational tools for structure determination.

DOI 10.1016/j.sbi.2008.06.009 - 2008

PubReading [197] - Nanopore microscope identifies RNA isoforms with structural colours - F. Bošković and U. Keyser

Wed, 12 Oct 2022 11:00:36 GMT

Identifying RNA transcript isoforms requires intricate protocols that suffer from various enzymatic biases. Here we design three-dimensional molecular constructs that enable identification of transcript isoforms at the single-molecule level using solid-state nanopore microscopy. We refold target RNA into RNA identifiers with designed sets of complementary DNA strands. Each reshaped molecule carries a unique sequence of structural (pseudo)colours. Structural colours consist of DNA structures, protein labels, native RNA structures or a combination of all three. The sequence of structural colours of RNA identifiers enables simultaneous identification and relative quantification of multiple RNA targets without prior amplification. Our Amplification-free RNA TargEt Multiplex Isoform Sensing (ARTEMIS) method reveals structural arrangements in native transcripts in agreement with published variants. ARTEMIS discriminates circular and linear transcript isoforms in a one-step, enzyme-free reaction in a complex human transcriptome using single-molecule read-out.

https://doi.org/10.1038/s41557-022-01037-5 -2022

PubReading [196] - Radiation damage to biological samples: still a pertinent issue - E. Garman and M. Weik

Tue, 11 Oct 2022 20:00:19 GMT

An understanding of radiation damage effects suffered by biological samples during structural analysis using both X-rays and electrons is pivotal to obtain reliable molecular models of imaged molecules. This special issue on radiation damage contains six papers reporting analyses of damage from a range of biophysical imaging techniques. For X-ray diffraction, an in-depth study of multi-crystal small-wedge data collection single-wavelength anomalous dispersion phasing protocols is presented, concluding that an absorbed dose of 5 MGy per crystal was optimal to allow reliable phasing. For small-angle X-ray scattering, experiments are reported that evaluate the efficacy of three radical scavengers using a protein designed to give a clear signature of damage in the form of a large conformational change upon the breakage of a disulfide bond. The use of X-rays to induce OH radicals from the radiolysis of water for X-ray footprinting are covered in two papers. In the first, new developments and the data collection pipeline at the NSLS-II high-throughput dedicated synchrotron beamline are described, and, in the second, the X-ray induced changes in three different proteins under aerobic and low-oxygen conditions are investigated and correlated with the absorbed dose. Studies in XFEL science are represented by a report on simulations of ultrafast dynamics in protic ionic liquids, and, lastly, a broad coverage of possible methods for dose efficiency improvement in modalities using electrons is presented.

doi.org/10.1107/S1600577521008845 - 2021

PubReading [195] - Inflammatory and immune effects on tumor progression - K. Mehla and M. Hollingsworth

Mon, 10 Oct 2022 20:00:17 GMT

Tumor progression is known occur in a complex microenvironment that leads to genetic, cellular, and metabolic adaptations. Two articles from Martin et al. and Del Poggetto et al. enlighten us on the role of inflammation and the immune system in guiding the progression of preneoplastic cells to oncogenic transformation and on subsequent tumor evolution.

doi.org/10.1016/j.it.2021.11.009 - 2022

PubReading [194] - Biological functions of therapy-induced senescence in cancer - E. Fitsiou, A. Soto-Gamez and M. Demaria

Fri, 07 Oct 2022 05:00:26 GMT

Therapy-induced cellular senescence is a state of stable growth arrest induced by common cancer treatments such as chemotherapy and radiation. In an oncogenic context, therapy-induced senescence can have different consequences. By blocking cellular proliferation and by facilitating immune cell infiltration, it functions as tumor suppressive mechanism. By fueling the proliferation of bystander cells and facilitating metastasis, it acts as a tumor promoting factor. This dual role is mainly attributed to the differential expression and secretion of a set of pro-inflammatory cytokines and tissue remodeling factors, collectively known as the Senescence-Associated Secretory Phenotype (SASP). Here, we describe cell-autonomous and non-cell-autonomous mechanisms that senescent cells activate in response to chemotherapy and radiation leading to tumor suppression and tumor promotion. We present the current state of knowledge on the stimuli that affect the activation of these opposing mechanisms and the effect of senescent cells on their micro-environment eg. by regulating the functions of immune cells in tumor clearance as well as strategies to eliminate senescent tumor cells before exerting their deleterious side-effects.

doi.org/10.1016/j.semcancer.2021.03.021 - 2021

PubReading [193] - A crystallization apparatus for temperature-controlled flow-cell dialysis with real-time visualization - N. Junius, M. Budayova-Spano et al.

Thu, 06 Oct 2022 20:00:18 GMT

Many instrumentation developments in crystallization have concentrated on massive parallelization assays and reduction of sample volume per experiment to find initial crystallization conditions. Yet improving the size and diffraction quality of the crystals for diffraction studies often requires decoupling of crystal nucleation and growth. This in turn requires the control of variables such as precipitant and protein concentration, equilibration rate, and temperature, which are all difficult parameters to control in the existing setups. The success of the temperature-controlled batch method, originally developed to grow very large crystals for neutron crystallography, demonstrated that the rational optimization of crystal growth has potential in structural biology. A temperature-controlled dialysis button has been developed for our previous device, and a prototype of an integrated apparatus for the rational optimization of crystal growth by mapping and manipulating temperature-precipitant concentration phase diagrams has been constructed. The presented approach differs from the current paradigm, since it involves serial instead of parallel experiments, exploring multiple crystallization conditions with the same protein sample. The sample is not consumed in the experiment and the conditions can be changed in a reversible fashion, using dialysis with a flowing precipitant reservoir as well as precise temperature control. The control software allows visualization of the crystals, as well as control of the temperature and composition of the crystallization solution. The rational crystallization optimization strategies presented here allow tailoring of crystal size, morphology and diffraction quality, significantly reducing the time, effort and amount of expensive protein material required for structure determination.

DOI: 10.1107/S1600576716004635 - 2016

PubReading [192] - Optimizing data collection for structure determination - A. GonzaÂlez

Wed, 05 Oct 2022 20:00:18 GMT

The ultimate purpose of diffraction data collection is to produce a data set which will result in the required structural information about the molecule of interest. This usually entails collecting a complete and accurate set of re ̄ection intensities to as high a resolution as possible. In practice, the characteristics of the crystal and properties of the X-ray source can be limiting factors to the data-set quality that can be achieved and a careful strategy has to be used to extract the maximum amount of information from the data within the experimental constraints. In the particular case of data intended for phasing using anomalous dispersion, the synchrotron beamline properties are relevant to determine how many wavelengths (one or more) should be used for the experiment and what the wavelength values should be. This will in turn affect the detailed strategy for data collection, including decisions about the data-collection sequence and how much data to collect at each wavelength. Collection of multiwavelength anomalous dispersion (MAD) data at three different wavelengths can provide very accurate experimental phases. Two-wavelength MAD experiments may offer the best compromise between phase quality and minimizing the effects of radiation damage to the sample. However, MAD experiments are demanding in terms of beamline wavelength range, easy tunability, stability and reproducibility. When the beamline cannot fulfill these demands, single-wavelength experiments may be a better option.

doi.org/10.1107/S0907444903017700 -2003

PubReading [191] - Review on mechanistic strategy of gene therapy in the treatment of disease - S. Alnasser

Tue, 04 Oct 2022 20:00:17 GMT

Gene therapy has become a revolution and its breakthrough is a corner stone in modern science. This treatment has rising advantages with limited negative aspects. Gene therapy is a therapeutic method in which, transfer of DNA to an individual to manipulate a defective gene is performed and to mitigate a disease which is not responding to pharmacological therapy. The gene therapy strategies are divided into two main categories such as direct in-vivo gene delivery of manipulated viral vector vehicle into the host and ex-vivo genetically engineered stem cells. In this review, we tried to cover all aspects of gene therapy studies; starting with the concept of gene, its treatment, gene delivery system and types, clinical trial either by vitro or In-Vivo -Clinical Trials and Clinical Intoxication of Gene Therapy. Therefore, the promise of successful treatment with gene therapy could positively affect millions of lives. The main aim of this review is to address the principles of gene therapy, various methods involved in the gene therapy, clinical applications and its merits and demerits

doi.org/10.1016/j.gene.2020.145246 - 2022

PubReading [189] - Clinical Experience With Gene Therapy in Older Patients With Spinal Muscular Atrophy - S. Metesanz, E. Kichula et al.

Fri, 30 Sep 2022 20:00:18 GMT

Onasemnogene abeparvovec was recently approved for the treatment of spinal muscular atrophy (SMA) in children younger than two years; however, clinical trials were primarily completed in children younger than seven months, so practical experience dosing older children began in summer 2019. Here, we look at the safety and efficacy of onasemnogene in seven infants older than seven months who were treated at our center. Overall, onasemnogene appears to be efficacious in children older than seven months and well tolerated. Side effects were similar to those previously reported, although more common and in some cases more severe and more prolonged than seen in the original trials. The impact of age, weight, and other confounding factors on development of side effects still needs to be elucidated.

doi.org/10.1016/j.pediatrneurol.2021.01.012 - 2021

PubReading [188] - m1A and m6A modifications function cooperatively to facilitate rapid mRNA degradation - S. Ho Boo, H. Ha, Y. Ki Kim

Thu, 29 Sep 2022 12:54:54 GMT

N6-Methyladenosine (m6A), the most abundant internal mRNA modification, affects multiple steps in gene expression. Mechanistically, the binding of YTHDF2 to m6A on mRNAs elicits rapid mRNA degradation by recruiting several RNA degrading enzymes. Here, we show that N1-methyladenosine (m1A), another type of RNA modification, accelerates rapid m6A RNA degradation. We identify HRSP12 as an RNA-binding protein that recognizes m1A. The binding of HRSP12 to m1A promotes efficient interaction of YTHDF2 with m6A, consequently facilitating endoribonucleolytic cleavage via the RNase P/MRP complex. Transcriptome- wide analyses also reveal that mRNAs harboring both m1A and m6A are downregulated in an HRSP12-dependent manner compared with mRNAs harboring m6A only. Accordingly, a subset of endogenous circular RNAs that harbor m6A and associate with YTHDF2 in an HRSP12-dependent manner is also subjected to m1A-facilitated rapid degradation. Together, our observations provide compelling evidence for crosstalk between different RNA modifications.

doi.org/10.1016/j.celrep.2022.111317 - 2022

PubReading [187] - T-cell invigoration to tumour burden ratio associated with anti-PD-1 response - A. Huang, J. Wherry et al

Tue, 27 Sep 2022 20:00:19 GMT

Despite the success of monotherapies based on blockade of programmed cell death 1 (PD-1) in human melanoma, most patients do not experience durable clinical benefit. Pre-existing T-cell infiltration and/or the presence of PD-L1 in tumours may be used as indicators of clinical response; however, blood-based profiling to understand the mechanisms of PD-1 blockade has not been widely explored. Here we use immune profiling of peripheral blood from patients with stage IV melanoma before and after treatment with the PD-1-targeting antibody pembrolizumab and identify pharmacodynamic changes in circulating exhausted-phenotype CD8 T cells (Tex cells). Most of the patients demonstrated an immunological response to pembrolizumab. Clinical failure in many patients was not solely due to an inability to induce immune reinvigoration, but rather resulted from an imbalance between T-cell reinvigoration and tumour burden. The magnitude of reinvigoration of circulating Tex cells determined in relation to pretreatment tumour burden correlated with clinical response. By focused profiling of a mechanistically relevant circulating T-cell subpopulation calibrated to pretreatment disease burden, we identify a clinically accessible potential on-treatment predictor of response to PD-1 blockade.

doi:10.1038/nature22079 - 2017

PubReading [186] - Copper(II) and silver(I)‐1,10‐phenanthroline‐5,6‐dione complexes interact with double‐stranded DNA: further evidence of their apparent multi‐modal activity towards Pseudomonas aeruginosa - A. Galdino, A. los Santos et al.

Mon, 26 Sep 2022 20:00:19 GMT

Tackling microbial resistance requires continuous efforts for the development of new molecules with novel mechanisms of action and potent antimicrobial activity. Our group has previously identified metal-based compounds, [Ag(1,10-phenan- throline-5,6-dione)2]ClO4 (Ag-phendione) and [Cu(1,10-phenanthroline-5,6-dione)3](ClO4)2.4H2O (Cu-phendione), with efficient antimicrobial action against multidrug-resistant species. Herein, we investigated the ability of Ag-phendione and Cu-phendione to bind with double-stranded DNA using a combination of in silico and in vitro approaches. Molecular dock- ing revealed that both phendione derivatives can interact with the DNA by hydrogen bonding, hydrophobic and electrostatic interactions. Cu-phendione exhibited the highest binding affinity to either major (− 7.9 kcal/mol) or minor (− 7.2 kcal/ mol) DNA grooves. In vitro competitive quenching assays involving duplex DNA with Hoechst 33258 or ethidium bromide demonstrated that Ag-phendione and Cu-phendione preferentially bind DNA in the minor grooves. The competitive eth- idium bromide displacement technique revealed Cu-phendione has a higher binding affinity to DNA (Kapp = 2.55 × 106 M−1) than Ag-phendione (Kapp = 2.79 × 105 M−1) and phendione (Kapp = 1.33 × 105 M−1). Cu-phendione induced topoisomerase I-mediated DNA relaxation of supercoiled plasmid DNA. Moreover, Cu-phendione was able to induce oxidative DNA inju- ries with the addition of free radical scavengers inhibiting DNA damage. Ag-phendione and Cu-phendione avidly displaced propidium iodide bound to DNA in permeabilized Pseudomonas aeruginosa cells in a dose-dependent manner as judged by flow cytometry. The treatment of P. aeruginosa with bactericidal concentrations of Cu-phendione (15 μM) induced DNA fragmentation as visualized by either agarose gel or TUNEL assays. Altogether, these results highlight a possible novel DNA-targeted mechanism by which phendione-containing complexes, in part, elicit toxicity toward the multidrug-resistant pathogen P. aeruginosa.

https://doi.org/10.1007/s00775-021-01922-3 - 2022

PubReading [185] - A methodology and an instrument for the temperature-controlled optimization of crystal growth - M. Budayova-Spano, S. Cusak et al

Fri, 23 Sep 2022 20:00:17 GMT

A method and a device for the promotion of crystal growth by keeping the crystallization solution metastable during the growth process are described. This is achieved by controlled temperature variation of the crystallization solution using parameters determined in situ during the growth process. The technique finds application in the growth of large high-quality crystals for neutron crystallography. Thus, it has been applied to grow large crystals of several proteins of interest such as human gamma-crystallin E, PA-IIL lectin from Pseudomonas aeruginosa, yeast inorganic pyrophosphatase, urate oxidase from Aspergillus flavus and human carbonic anhydrase II.

doi:10.1107/S0907444906054230 - 2007

PubReading [184] - Biophysical Techniques in Structural Biology - C. Dobson

Thu, 22 Sep 2022 20:00:19 GMT

Over the past six decades, steadily increasing progress in the application of the principles and techniques of the physical sciences to the study of biological systems has led to remarkable insights into the molecular basis of life. Of particular significance has been the way in which the determination of the structures and dynamical properties of proteins and nucleic acids has so often led directly to a profound understanding of the nature and mechanism of their functional roles. The increasing number and power of experimental and theoretical techniques that can be applied successfully to living systems is now ushering in a new era of structural biology that is leading to fundamentally new information about the maintenance of health, the origins of disease, and the development of effective strategies for therapeutic intervention. This article provides a brief overview of some of the most powerful biophysical methods in use today, along with references that provide more detailed information about recent applications of each of them. In addition, this article acts as an introduction to four authoritative reviews in this volume. The first shows the ways that a multiplicity of bio-physical methods can be combined with computational techniques to define the architectures of complex biological systems, such as those involving weak interactions within ensembles of molecular components. The second illustrates one aspect of this general approach by describing how recent advances in mass spectrometry, particularly in combination with other techniques, can generate fundamentally new insights into the properties of membrane proteins and their functional interactions with lipid molecules. The third review demonstrates the increasing power of rapidly evolving diffraction techniques, employing the very short bursts of X-rays of extremely high intensity that are now accessible as a result of the construction of free-electron lasers, in particular to carry out time-resolved studies of biochemical reactions. The fourth describes in detail the application of such approaches to probe the mechanism of the light-induced changes associated with bacteriorhodopsin’s ability to convert light energy into chemical energy.

doi.org/10.1146/annurev-biochem-013118- 111947 - 2019

PubReading [183] - From structure to function: Route to understanding lncRNA mechanism - J. Graf and M. Kretz

Wed, 21 Sep 2022 20:00:19 GMT

RNAs have emerged as a major target for diagnostics and therapeutics approaches. Regulatory nonprotein-coding RNAs (ncRNAs) in particular display remarkable ver- satility. They can fold into complex structures and interact with proteins, DNA, and other RNAs, thus modulating activity, localization, or interactome of multi-protein complexes. Thus, ncRNAs confer regulatory plasticity and represent a new layer of reg- ulatory control. Interestingly, long noncoding RNAs (lncRNAs) tend to acquire complex secondary and tertiary structures and their function—in many cases—is dependent on structural conservation rather than primary sequence conservation. Whereas for many proteins, structure and its associated function are closely con- nected, for lncRNAs, the structural domains that determine functionality and its inter- actome are still not well understood. Numerous approaches for analyzing the struc- tural configuration of lncRNAs have been developed recently. Here, will provide an overview of major experimental approaches used in the field, and discuss the poten- tial benefit of using combinatorial strategies to analyze lncRNA modes of action based on structural information.

DOI: 10.1002/bies.202000027 - 2020

PubReading [182] - Three's a crowd – stabilisation, structure, and applications of DNA triplexes - M. Dalla Pozza, A. Abdullrahman, C. Cardin, G. Gasser and J. Hall

Tue, 20 Sep 2022 20:00:18 GMT

DNA is a strikingly flexible molecule and can form a variety of secondary structures, including the triple helix, which is the subject of this review. The DNA triplex may be formed naturally, during homologous recombination, or can be formed by the introduction of a synthetic triplex forming oligonucleotide (TFO) to a DNA duplex. As the TFO will bind to the duplex with sequence specificity, there is significant interest in developing TFOs with potential therapeutic applications, including using TFOs as a delivery mechanism for compounds able to modify or damage DNA. However, to combine triplexes with functionalised compounds, a full understanding of triplex structure and chemical modification strategies, which may increase triplex stability or in vivo degradation, is essential – these areas will be discussed in this review. Ruthenium polypyridyl complexes, which are able to photooxidise DNA and act as luminescent DNA probes, may serve as a suitable photophysical payload for a TFO system and the developments in this area in the context of DNA triplexes will also be reviewed.

DOI: 10.1039/d2sc01793h - 2022

PubReading [181] - Degraders: The Ultimate Weapon Against Amplified Driver Kinases in Cancer - P. Torres-Ayuso and J. Brognard

Mon, 19 Sep 2022 20:00:18 GMT

Amplification of pro-oncogenic kinases is a common genetic alteration driving tumorigenic phenotypes. Cancer cells rely on the amplified kinases to sustain cell proliferation, survival, and growth, presenting an opportunity to develop therapies targeting the amplified kinases. Utilizing small molecule catalytic inhibitors as therapies to target amplified kinases is plagued by de novo resistance driven by increased expression of the target, and amplified kinases can drive tumorigenic phenotypes independent of catalytic activity. Here, we discuss the emergence of proteolysis-targeting chimeras that provide an opportunity to target these oncogenic drivers effectively.

doi.org/10.1124/molpharm.121.000306 - 2022

PubReading [180] - Glioblastoma hijacks neuronal mechanisms for brain invasion - V. Venkataramani, F. Winkler et al.

Fri, 16 Sep 2022 20:00:30 GMT

Glioblastomas are incurable tumors infiltrating the brain. A subpopulation of glioblastoma cells forms a functional and therapy-resistant tumor cell network interconnected by tumor microtubes (TMs). Other sub-populations appear unconnected, and their biological role remains unclear. Here, we demonstrate that whole-brain colonization is fueled by glioblastoma cells that lack connections with other tumor cells and astrocytes yet receive synaptic input from neurons. This subpopulation corresponds to neuronal and neural-progenitor-like tumor cell states, as defined by single-cell transcriptomics, both in mouse models and in the human disease. Tumor cell invasion resembled neuronal migration mechanisms and adopted a Levy-like movement pattern of probing the environment. Neuronal activity induced complex calcium signals in glioblastoma cells followed by the de novo formation of TMs and increased invasion speed. Collectively, superimposing molecular and functional single-cell data revealed that neuronal mechanisms govern glioblastoma cell invasion on multiple levels. This explains how glioblastoma’s dissemination and cellular heterogeneity are closely interlinked.

doi.org/10.1016/j.cell.2022.06.054 - 2022

PubReading [179] - Metallodrugs: Mechanisms of Action, Molecular Targets and Biological Activity - G. Ferrano and A. Merlino

Thu, 15 Sep 2022 20:00:20 GMT

The research interest in the field of inorganic medicinal chemistry had a large increase after the serendipitous discovery of the cytotoxic activity of cisplatin by Rosenberg at the end of 1960s. Since then, cisplatin has entered clinical practice and become one of the most common treatments for solid tumors. Unfortunately, the use of cisplatin and its derivatives is associated with undesired side effects, such as general toxicity and intrinsic and acquired drug-resistance. For these reasons, alternative metallodrugs based on non- Pt metals have been synthetized, characterized and tested for biological activity in recent years. Although, initially, DNA seemed the exclusive target for metallodrugs [6], successive studies have revealed that various metabolites, peptides and proteins have a central role in the recognition, transport and mechanism of action of these compounds. Thus, a deeper understanding of the molecular bases of the interaction of metallodrugs with these molecules is needed.

doi.org/10.3390/ijms23073504 - 2022

PubReading [178] - Mass spectrometry techniques for imaging and detection of metallodrugs - S. Theiner, G. Koellensperger et al.

Wed, 14 Sep 2022 20:00:16 GMT

Undoubtedly, metallomic approaches based on mass spectrometry have evolved into essential tools supporting the drug development of novel metal-based anticancer drugs. This article will comment on the state-of-the-art instrumentation and highlight some of the recent analytical advances beyond routine, especially focusing on the latest developments in inductively coupled plasma-mass spectrometry (ICP-MS). Mass spectrometry-based bioimaging and single-cell methods will be presented, paving the way to exciting investigations of metal-based anticancer drugs in heterogeneous and structurally, as well as functionally complex solid tumor tissues.

doi.org/10.1016/j.cbpa.2020.12.005 - 2021

PubReading [177] - Systematic Review of miRNA as Biomarkers in Alzheimer’s Disease - S. Swarbrick, N. Wragg, S. Ghosh and A. Stolzing

Tue, 13 Sep 2022 20:00:17 GMT

Currently there are 850,000 people with Alzheimer’s disease in the UK, with an estimated rise to 1.1 million by 2025. Alzheimer’s disease is characterised by the accumulation of amyloid-beta plaques and hyperphosphorylated tau in the brain causing a progressive decline in cognitive impairment. Small non-coding microRNA (miRNA) sequences have been found to be deregulated in the peripheral blood of Alzheimer patients. A systematic review was conducted to extract all miRNA found to be significantly deregulated in the peripheral blood. These deregulated miRNAs were cross-referenced against the miRNAs deregulated in the brain at Braak Stage III. This resulted in a panel of 10 miRNAs (hsa-mir-107, hsa-mir-26b, hsa-mir-30e, hsa-mir-34a, hsa-mir-485, hsa-mir200c, hsa-mir-210, hsa-mir-146a, hsa-mir-34c, and hsa-mir-125b) hypothesised to be deregulated early in Alzheimer’s disease, nearly 20 years before the onset of clinical symptoms. After network analysis of the 10 miRNAs, they were found to be associated with the immune system, cell cycle, gene expression, cellular response to stress, neuron growth factor signalling, wnt signalling, cellular senescence, and Rho GTPases.

doi.org/10.1007/s12035-019-1500-y - 2019

PubReading [176] - miRNA Targeting: Growing beyond the Seed - L. Chipman and A. Pasquinelli

Mon, 12 Sep 2022 20:00:29 GMT

miRNAs are small RNAs that guide Argonaute proteins to specific target mRNAs to repress their translation and stability. Canonically, miRNA targeting is reliant on base pairing of the seed region, nucleotides 2–7, of the miRNA to sites in mRNA 30 untranslated regions. Recently, the 30 half of the miRNA has gained attention for newly appreciated roles in regulating target specificity and regulation. In addition, the extent of pairing to the miRNA 30 end can influence the stability of the miRNA itself. These findings highlight the importance of sequences beyond the seed in controlling the function and existence of miRNAs.

doi.org/10.1016/j.tig.2018.12.005 - 2018

PubReading [175] - Solution Oligonucleotide APIs: Regulatory Considerations - C. Wetter. J. Tom et al.

Fri, 09 Sep 2022 20:00:19 GMT

Manufacture of oligonucleotide active pharmaceutical ingredients (APIs) typically consists of solid-phase synthesis, deprotection and cleavage, purification and filtration, and isolation from aqueous solutions through lyophilization. In the first step of drug product manufacture, the API is dissolved in water again and excipients are added. While isolation of oligonucleotide APIs can be meaningful in many cases, there may be cases where keeping the API in solution provides benefit, and multiple technical aspects must be taken into account and balanced when determining the appropriate API form. A significant factor is whether an API in solution will contain additional components. While APIs in solution containing additional components (so-called formulated APIs) are well established for biological products, there are regulatory guidelines in place that represent hurdles for industry to using a formulated API approach for oligonucleotide drugs. The present communication outlines conditions where a formulated API approach can be chosen in compliance with existing guidelines. Relevant aspects pertaining to risk management, GMP standards, facility design, control strategies, and regulatory submission content are discussed. In addition, the authors propose that existing guidelines be modernized to enable the use of a formulated API approach for additional reasons than the ones described in the existing regulatory framework. The manuscript aims to promote a dialog with regulators in this field.

doi.org/10.1007/s43441-022-00384-2 - 2022

PubReading [174] - All-atom simulations to studying metallodrugs/target interactions - P. Janos, A. Spinello and A. Magistrato

Thu, 08 Sep 2022 20:00:20 GMT

Metallodrugs are extensively used to treat and diagnose distinct disease types. The unique physical-chemical properties of metal ions offer tantalizing opportunities to tailor effective scaffolds for selectively targeting specific biomolecules. Modern experimental techniques have collected a large body of structural data concerning the interactions of metallodrugs with their biomolecular targets, although being unable to exhaustively assess the molecular basis of their mechanism of action. In this scenario, the complementary use of accurate computational methods allows uncovering the minutiae of metallodrugs/targets interactions and their underlying mechanism of action at an atomic level of detail. This knowledge is increasingly perceived as an invaluable requirement to rationally devise novel and selective metallodrugs. Building on literature studies, selected largely from the last 2 years, this compendium encompasses a cross-section of the current role, advances, and challenges met by computer simulations to decipher the mechanistic intricacies of prototypical metallodrugs.

doi.org/10.1016/j.cbpa.2020.07.005 -2020

PubReading [173] - CAR T cell immunotherapy for human cancer - C. June, M. Milone et al.

Wed, 07 Sep 2022 20:00:19 GMT

Adoptive T cell transfer (ACT) is a new area of transfusion medicine involving the infusion of lymphocytes to mediate antitumor, antiviral, or anti-inflammatory effects. The field has rapidly advanced from a promising form of immuno-oncology in preclinical models to the recent commercial approvals of chimeric antigen receptor (CAR) T cells to treat leukemia and lymphoma. This Review describes opportunities and challenges for entering mainstream oncology that presently face the CAR T field, with a focus on the challenges that have emerged over the past several years.

DOI: 10.1126/science.aar6711 - 2018

PubReading [172] - Delivery of oligonucleotide-based therapeutics: challenges and opportunities - S. Hammond, V. Arechavala-Gomeza et.

Tue, 06 Sep 2022 20:00:19 GMT

Nucleic acid-based therapeutics that regulate gene expression have been developed towards clinical use at a steady pace for several decades, but in recent years the field has been accelerating. To date, there are 11 marketed products based on antisense oligonucleotides, aptamers and small interfering RNAs, and many others are in the pipeline for both academia and industry. A major technology trigger for this development has been progress in oligonucleotide chemistry to improve the drug properties and reduce cost of goods, but the main hurdle for the application to a wider range of disorders is delivery to target tissues. The adoption of delivery technologies, such as conjugates or nanoparticles, has been a game changer for many therapeutic indications, but many others are still awaiting their eureka moment. Here, we cover the variety of methods developed to deliver nucleic acid-based therapeutics across biological barriers and the model systems used to test them. We discuss important safety considerations and regulatory requirements for synthetic oligonucleotide chemistries and the hurdles for translating laboratory breakthroughs to the clinic. Recent advances in the delivery of nucleic acid-based therapeutics and in the development of model systems, as well as safety considerations and regulatory requirements for synthetic oligonucleotide chemistries are discussed in this review on oligonucleotide-based therapeutics.

DOI 10.15252/emmm.202013243 - 2020

PubReading [171] - Dual action of ketamine confines addiction liability - L. Simmler, C. Lüscher et al.

Mon, 05 Sep 2022 20:00:17 GMT

Ketamine is used clinically as an anaesthetic and a fast-acting antidepressant, and recreationally for its dissociative properties, raising concerns of addiction as a possible side effect. Addictive drugs such as cocaine increase the levels of dopamine in the nucleus accumbens. This facilitates synaptic plasticity in the mesolimbic system, which causes behavioural adaptations and eventually drives the transition to compulsion1–4. The addiction liability of ketamine is a matter of much debate, in part because of its complex pharmacology that among several targets includes N-methyl-d-aspartic acid (NMDA) receptor (NMDAR) antagonism5,6. Here we show that ketamine does not induce the synaptic plasticity that is typically observed with addictive drugs in mice, despite eliciting robust dopamine transients in the nucleus accumbens. Ketamine nevertheless supported reinforcement through the disinhibition of dopamine neurons in the ventral tegmental area (VTA). This effect was mediated by NMDAR antagonism in GABA (γ-aminobutyric acid) neurons of the VTA, but was quickly terminated by type-2 dopamine receptors on dopamine neurons. The rapid off-kinetics of the dopamine transients along with the NMDAR antagonism precluded the induction of synaptic plasticity in the VTA and the nucleus accumbens, and did not elicit locomotor sensitization or uncontrolled self-administration. In summary, the dual action of ketamine leads to a unique constellation of dopamine-driven positive reinforcement, but low addiction liability.

doi.org/10.1038/s41586-022-04993-7 - 2022

PubReading [170] - Accelerating reaction generality and mechanistic insight through additive mapping - C. Kullmer, D. MacMillan et al

Fri, 02 Sep 2022 20:00:17 GMT

Reaction generality is crucial in determining the overall impact and usefulness of synthetic methods. Typical generalization protocols require a priori mechanistic understanding and suffer when applied to complex, less understood systems. We developed an additive mapping approach that rapidly expands the utility of synthetic methods while generating concurrent mechanistic insight. Validation of this approach on the metallaphotoredox decarboxylative arylation resulted in the discovery of a phthalimide ligand additive that overcomes many lingering limitations of this reaction and has important mechanistic implications for nickel-catalyzed cross-couplings.

DOI: 10.1126/science.abn1885 - 2022

PubReading [169] - The Epigenetic Basis of Hematopoietic Stem Cell Aging - A. Kramer and G. Challen

Thu, 01 Sep 2022 20:00:20 GMT

Highly proliferative tissues such as the gut, skin and bone marrow lose millions of cells each day to normal attrition and challenge from different biological adversities. To achieve a lifespan beyond the longevity of individual cell types, tissue-specific stem cells sustain these tissues throughout the life of a human. For example, the lifespan of erythrocytes is about 100 days and adults make about two million new erythrocytes every second. A small pool of hematopoietic stem cells (HSCs) in the bone marrow is responsible for the lifetime maintenance of these populations. However, there are changes that occur within the HSC pool during aging. Biologically, these changes manifest as blunted immune responses, decreased bone marrow cellularity, and increased risk of myeloid diseases. Understanding the molecular mechanisms underlying dysfunction of aging HSCs is an important focus of biomedical research. With advances in modern health care, the average age of the general population is ever increasing. If molecular or pharmacological interventions could be discovered that rejuvenate aging HSCs, it could reduce the burden of age related immune system compromise as well as open up new opportunities for treatment of hematological disorders with regenerative therapy.

doi:10.1053/j.seminhematol.2016.10.006 - 2017

PubReading [168] - Advances of epigenetic editing - R. Gjaltema and M. Rots

Fri, 29 Jul 2022 20:00:19 GMT

Epigenetic editing refers to the locus-specific targeting of epigenetic enzymes to rewrite the local epigenetic landscape of an endogenous genomic site, often with the aim of transcriptional reprogramming. Implementing clustered regularly interspaced short palindromic repeat–dCas9 greatly accelerated the advancement of epigenetic editing, yielding preclinical therapeutic successes using a variety of epigenetic enzymes. Here, we review the current applications of these epigenetic editing tools in mammals and shed light on biochemical improvements that facilitate versatile applications.

doi.org/10.1016/j.cbpa.2020.04.020 - 2020

PubReading [167] - Molecular Biology and Evolution of Cancer: From Discovery to Action - J. Somarelli, J. Townsend et al

Thu, 28 Jul 2022 20:00:18 GMT

Cancer progression is an evolutionary process. During this process, evolving cancer cell populations encounter restrictive ecological niches within the body, such as the primary tumor, circulatory system, and diverse metastatic sites. Efforts to prevent or delay cancer evolution—and progression—require a deep understanding of the underlying molecular evolutionary processes. Herein we discuss a suite of concepts and tools from evolutionary and ecological theory that can inform cancer biology in new and meaningful ways. We also highlight current challenges to applying these concepts, and propose ways in which incorporating these concepts could identify new therapeutic modes and vulnerabilities in cancer.

doi:10.1093/molbev/msz242 - 2019

PubReading [166] - Understanding the fate of DNA nanostructures inside the cell - C. Green, D. Mathur and I. Medinitz

Wed, 27 Jul 2022 20:00:26 GMT

Structural DNA nanotechnology is poised to transform targeted therapeutic and theranostic delivery agents. Some of the most promising biomedical applications of DNA nanostructures include carriers for biosensing, imaging, and drug delivery. Additionally, the unique ability to precisely position inorganic and organic molecules on DNA-based substrates enables the spatially optimized high density interfacing of ligands with cell membrane receptors. To realize clinically viable biomedical products made from DNA nanostructures, it is necessary to fully understand the behavior of these systems inside and outside the cellular environment. To that end, cohesive and conclusive information on the physiological fate of DNA nanostructures at various time points – from the cell culture to the cell cytosol – is still lacking. In this highlight, we bring to attention efforts to understand DNA nanostructure behavior in vitro as well as some widespread disparities among studies on the subject. We also call for a discussion on the implementation of common standards and controls to address these disparities and consequently unify the scientific community’s endeavours to build foundational knowledge on DNA nanostructure–cellular interaction.

DOI: 10.1039/d0tb00395f - 2020

PubReading [165] - Macrocyclic Metal Complex−DNA Conjugates for Electrochemical Sensing of Single Nucleobase Changes in DNA - J. Duprey, J. Tucker et al

Tue, 26 Jul 2022 20:00:19 GMT

The direct incorporation of macrocyclic cyclidene complexes into DNA via automated synthesis results in a new family of metal-functionalized DNA derivatives that readily demonstrate their utility through the ability of one redox-active copper(II)-containing strand to distinguish electrochemically between all four canonical DNA nucleobases at a single site within a target sequence of DNA.

DOI: 10.1021/jacs.5b11319 - 2015

PubReading [164] - DNA Nanostructures for Targeted Antimicrobial Delivery - I. Mela, C. Kaminiski et al

Mon, 25 Jul 2022 20:00:18 GMT

We report the use of DNA origami nanostructures, functionalized with aptamers, as a vehicle for delivering the antibacterial enzyme lysozyme in a specific and efficient manner. We test the system against Gram-positive (Bacillus subtilis) and Gram-negative (Escherichia coli) targets. We use direct stochastic optical reconstruction microscopy (dSTORM) and atomic force microscopy (AFM) to characterize the DNA origami nanostructures and structured illumination microscopy (SIM) to assess the binding of the origami to the bacteria. We show that treatment with lysozyme-functionalized origami slows bacterial growth more effectively than treatment with free lysozyme. Our study introduces DNA origami as a tool in the fight against antibiotic resistance, and our results demonstrate the specificity and efficiency of the nanostructure as a drug delivery vehicle.

doi.org/10.1002/anie.202002740 - 2020

PubReading [163] - Synthetic protein-conductive membrane nanopores built with DNA - T. Diederichs, R. Tampé, S. Howorka et al.

Fri, 08 Jul 2022 20:00:18 GMT

Nanopores are key in portable sequencing and research given their ability to transport elongated DNA or small bioactive molecules through narrow transmembrane channels. Transport of folded proteins could lead to similar scientific and technological benefits. Yet this has not been realised due to the shortage of wide and structurally defined natural pores. Here we report that a synthetic nanopore designed via DNA nanotechnology can accommodate folded proteins. Transport of fluorescent proteins through single pores is kinetically analysed using massively parallel optical readout with transparent silicon-on-insulator cavity chips vs. electrical recordings to reveal an at least 20-fold higher speed for the electrically driven movement. Pores nevertheless allow a high diffusive flux of more than 66 molecules per second that can also be directed beyond equillibria. The pores may be exploited to sense diagnostically relevant proteins with portable analysis technology, to create molecular gates for drug delivery, or to build synthetic cells.

doi.org/10.1038/s41467-019-12639-y - 2019

PubReading [162] - The second decade of synthetic biology: 2010–2020 - F. Meng and T. Ellis

Thu, 07 Jul 2022 20:00:18 GMT

Synthetic biology is among the most hyped research topics this century, and in 2010 it entered its teenage years. But rather than these being a problematic time, we’ve seen synthetic biology blossom and deliver many new technologies and landmark achievements.

doi.org/10.1038/s41467-020-19092-2 - 2020

PubReading [161] - Even Cancer Cells Watch Their Cholesterol! - R. Riscal, N. Skuli and C. Simon

Wed, 06 Jul 2022 20:00:18 GMT

Deregulated cell proliferation is an established feature of cancer, and altered tumor metabolism has witnessed renewed interest over the past decade, including the study of how cancer cells rewire metabolic pathways to renew energy sources and ‘‘building blocks’’ that sustain cell division. Microenvironmental oxygen, glucose, and glutamine are regarded as principal nutrients fueling tumor growth. However, hostile tumor microenvironments render O2/nutrient supplies chronically insufficient for increased proliferation rates, forcing cancer cells to develop strategies for opportunistic modes of nutrient acquisition. Recent work shows that cancer cells overcome this nutrient scarcity by scavenging other substrates, such as proteins and lipids, or utilizing adaptive metabolic pathways. As such, reprogramming lipid metabolism plays important roles in providing energy, macromolecules for membrane synthesis, and lipid-mediated signaling during cancer progression. In this review, we highlight more recently appreciated roles for lipids, particularly cholesterol and its derivatives, in cancer cell metabolism within intrinsically harsh tumor microenvironments.

doi.org/10.1016/j.molcel.2019.09.008 - 2019

PubReading [160] - Sequence-Selective Minor Groove Recognition of a DNA Duplex Containing Synthetic Genetic Components - G. Padroni, G. Burley et al.

Tue, 05 Jul 2022 20:00:19 GMT

The structural basis of minor groove recognition of a DNA duplex containing synthetic genetic information by hairpin pyrrole- imidazole polyamides is described. Hairpin polyamides induce a higher melting stabilization of a DNA duplex containing the unnatural P·Z base- pair when an imidazole unit is aligned with a P nucleotide. An NMR structural study showed that the incorporation of two isolated P·Z pairs enlarges the minor groove and slightly narrows the major groove at the site of this synthetic genetic information, relative to a DNA duplex consisting entirely of Watson−Crick base-pairs. Pyrrole-imidazole polyamides bind to a P·Z-containing DNA duplex to form a stable complex, effectively mimicking a G·C pair. A structural hallmark of minor groove recognition of a P·Z pair by a polyamide is the reduced level of allosteric distortion induced by binding of a polyamide to a DNA duplex. Understanding the molecular determinants that influence minor groove recognition of DNA containing synthetic genetic components provides the basis to further develop unnatural base-pairs for synthetic biology applications.

DOI: 10.1021/jacs.8b12444 - 2019

PubReading [159] - Designing Hydrogels for On-Demand Therapy - N. Oliva, N. Artzi et al.

Mon, 04 Jul 2022 20:00:18 GMT

Systemic administration of therapeutic agents has been the preferred approach to treat most pathological conditions, in particular for cancer therapy. This treatment modality is associated with side effects, off-target accumulation, toxicity, and rapid renal and hepatic clearance. Multiple efforts have focused on incorporating targeting moieties into systemic therapeutic vehicles to enhance retention and minimize clearance and side effects. However, only a small percentage of the nanoparticles administered systemically accumulate at the tumor site, leading to poor therapeutic efficacy. This has prompted researchers to call the status quo treatment regimen into question and to leverage new delivery materials and alternative administration routes to improve therapeutic outcomes. Recent approaches rely on the use of local delivery platforms that circumvent the hurdles of systemic delivery. Local administration allows delivery of higher “effective” doses while enhancing therapeutic molecules’ stability, minimizing side effects, clearance, and accumulation in the liver and kidneys following systemic administration. Hydrogels have proven to be highly biocompatible materials that allow for versatile design to afford sensing and therapy at the same time. Hydrogels’ chemical and physical versatility can be exploited to attain disease-triggered in situ assembly and hydrogel programmed degradation and consequent drug release, and hydrogels can also serve as a biocompatible depot for local delivery of stimuli-responsive therapeutic cargo. We will focus this Account on the hydrogel platform that we have developed in our lab, based on dendrimer amine and dextran aldehyde. This hydrogel is disease-responsive and capable of sensing the microenvironment and reacting in a graded manner to diverse pathologies to render different properties, including tissue adhesion, biocompatibility, hydrogel degradation, and embedded drug release profile. We also studied the degradation kinetics of our stimuli-responsive materials in vivo and analyzed the in vitro conditions under which in vitro−in vivo correlation is attained. Identifying key parameters in the in vivo microenvironment under healthy and disease conditions was key to attaining that correlation. The adhesive capacity of our dendrimer−dextran hydrogel makes it optimal for localized and sustained release of embedded drugs. We demonstrated that it affords the delivery of a range of therapeutics to combat cancer, including nucleic acids, small molecules, and antibody drugs. As a depot for local delivery, it allows a high dose of active biomolecules to be delivered directly at the tumor site. Immunotherapy, a recently blooming area in cancer therapy, may exploit stimuli-responsive hydrogels to impart systemic effects following localized therapy. Local delivery would enable release of the proper drug dose and improve drug bioavailability where needed at the same time creating memory and exerting the therapeutic effect systemically. This Account highlights our perspective on how local and systemic therapies provided by stimuli-responsive hydrogels should be used to impart more precise, long-lasting, and potent therapeutic outcomes.

DOI: 10.1021/acs.accounts.6b00536 - 2017

PubReading [158] - Junctions in DNA: underexplored targets for therapeutic intervention - E. Ivens, M. Cominetti and M. Searcey

Fri, 01 Jul 2022 20:00:18 GMT

DNA has been a key target for cancer therapy, with a range of compounds able to bind and either impair its processing or induce damage. Targeting DNA with small molecules in a truly sequence specific way, to impair gene specific processes, remains out of reach. The ability of DNA to assume different structures from the classical double helix allows access to more specific ligand binding modes and, potentially, to new avenues of treatment. In this review, we illustrate the small molecules that have been reported to bind to three- and four-way junctions.

doi.org/10.1016/j.bmc.2022.116897 - 2022

PubReading [157] - Cell-SELEX Technology - S. Ohuchi

Thu, 30 Jun 2022 20:00:18 GMT

Aptamers are molecules identified from large combinatorial nucleic acid libraries by their high affinity to target molecules. Due to a variety of desired properties, aptamers are attractive alternatives to antibodies in molecular biology and medical applications. Aptamers are identified through an iterative selection–amplification process known as systematic evolution of ligands by exponential enrichment (SELEX). Although SELEX is typically carried out using purified target molecules, whole live cells are also employable as selection targets. This technology, Cell-SELEX, has several advantages. For example, generated aptamers are functional with a native conformation of the target molecule on live cells, and thus, cell surface transmembrane proteins would be targets even went their purifications in native conformations are difficult. In addition, cell-specific aptamers can be obtained without any knowledge about cell surface molecules on the target cells. Here, I review the progress of Cell-SELEX technology and discuss advantages of the technology

DOI: 10.1089/biores.2012.0253265 - 2012

PubReading [156] - Direct Nanopore Sequencing of Individual Full Length tRNA Strands - N. Thomas, R. Abu-Shumays et al

Wed, 29 Jun 2022 20:00:17 GMT

We describe a method for direct tRNA sequencing using the Oxford Nanopore MinION. The principal technical advance is custom adapters that facilitate end-to-end sequencing of individual transfer RNA (tRNA) molecules at subnanometer precision. A second advance is a nanopore sequencing pipeline optimized for tRNA. We tested this method using purified E. coli tRNAfMet, tRNALys, and tRNAPhe samples. 76–92% of individual aligned tRNA sequence reads were full length. As a proof of concept, we showed that nanopore sequencing detected all 43 expected isoacceptors in total E. coli MRE600 tRNA as well as isodecoders that further define that tRNA population. Alignment-based comparisons between the three purified tRNAs and their synthetic controls revealed systematic nucleotide miscalls that were diagnostic of known modifications. Systematic miscalls were also observed proximal to known modifications in total E. coli tRNA alignments, including a highly conserved pseudouridine in the T loop. This work highlights the potential of nanopore direct tRNA sequencing as well as improvements needed to implement tRNA sequencing for human healthcare applications.

doi.org/10.1021/acsnano.1c06488 - 2021

PubReading [155] - The central role of DNA damage in the ageing process - B. Schumacher, J. Pothof, Vijg and Hoeijmakers

Tue, 28 Jun 2022 20:00:18 GMT

Ageing is a complex, multifaceted process leading to widespread functional decline that affects every organ and tissue, but it remains unknown whether ageing has a unifying causal mechanism or is grounded in multiple sources. Phenotypically, the ageing process is associated with a wide variety of features at the molecular, cellular and physiological level—for example, genomic and epigenomic alterations, loss of proteostasis, declining overall cellular and subcellular function and deregulation of signalling systems. However, the relative importance, mechanistic interrelationships and hierarchical order of these features of ageing have not been clarified. Here we synthesize accumulating evidence that DNA damage affects most, if not all, aspects of the ageing phenotype, making it a potentially unifying cause of ageing. Targeting DNA damage and its mechanistic links with the ageing phenotype will provide a logical rationale for developing unified interventions to counteract age-related dysfunction and disease.

doi.org/10.1038/s41586-021-03307-7 - 2021

PubReading [154] - Mitochondria‐ER Tethering in Neurodegenerative Diseases - R. Raeisossadati, M. Ferrari

Mon, 27 Jun 2022 20:00:19 GMT

Organelles juxtaposition has been detected for decades, although only recently gained importance due to a pivotal role in the regulation of cellular processes dependent on membrane contact sites. Endoplasmic reticulum (ER) and mitochondria interaction is a prime example of organelles contact sites. Mitochondria-associated membranes (MAM) are proposed to harbor ER-mitochondria tether complexes, mainly when these organelles are less than 30 nm apart. Dysfunctions of proteins located at the MAM are associated with neurodegenerative diseases such as Parkinson’s, Alzheimer’s and amyotrophic lateral sclerosis, as well as neurodevelopmental disorders; hence any malfunction in MAM can potentially trigger cell death. This review will focus on the role of ER-mitochondria contact sites, regarding calcium homeostasis, lipid metabolism, autophagy, morphology and dynamics of mitochondria, mainly in the context of neurodegenerative diseases. Approaches that have been employed so far to study organelles contact sites, as well as methods that were not used in neurosciences yet, but are promising and accurate ways to unveil the functions of MAM during neurodegeneration, is also discussed in the present review.

doi.org/10.1007/s10571-020-01008-9 - 2022

PubReading [153] - The DNA methyltransferase family: a versatile toolkit for epigenetic regulation - Frank Lyko

Fri, 24 Jun 2022 20:00:19 GMT

The DNA methyltransferase (DNMT) family comprises a conserved set of DNA-modifying enzymes that have a central role in epigenetic gene regulation. Recent studies have shown that the functions of the canonical DNMT enzymes — DNMT1, DNMT3A and DNMT3B — go beyond their traditional roles of establishing and maintaining DNA methylation patterns. This Review analyses how molecular interactions and changes in gene copy numbers modulate the activity of DNMTs in diverse gene regulatory functions, including transcriptional silencing, transcriptional activation and post-transcriptional regulation by DNMT2-dependent tRNA methylation. This mechanistic diversity enables the DNMT family to function as a versatile toolkit for epigenetic regulation.

doi:10.1038/nrg.2017.80 - 2018

PubReading [152] - Teaching photosensitizers a new trick: red light-triggered G-quadruplex alkylation by ligand co-localization - E. Cadoni, A. Madder et al

Thu, 23 Jun 2022 20:00:18 GMT

We propose a bimolecular approach for G-quadruplex alkylation, using a pro-reactive furan-containing ligand, activated by red-light irradiation of a proximate G4-binding photosensitizer. G4- over dsDNA alkylation can be achieved selectively and proves high-yielding at low ligand excess. HPLC and modelling studies allowed identifying potential residues involved in the alkylation.

DOI: 10.1039/d0cc06030e - 2021

PubReading [151] - Chirality transmission in macromolecular domains - S. Pandey, C. Lou, H.Mao et al

Wed, 22 Jun 2022 20:00:18 GMT

Chiral communications exist in secondary structures of foldamers and copolymers via a network of noncovalent interactions within effective intermolecular force (IMF) range. It is not known whether long-range chiral communication exists between macromolecular tertiary structures such as peptide coiled-coils beyond the IMF distance. Harnessing the high sensitivity of single-molecule force spectroscopy, we investigate the chiral interaction between covalently linked DNA duplexes and peptide coiled-coils by evaluating the binding of a diastereomeric pair of three DNA-peptide conjugates. We find that right-handed DNA triple helices well accommodate peptide triple coiled-coils of the same handedness, but not with the left-handed coiled-coil stereoisomers. This chiral communication is effective in a range (<4.5 nm) far beyond canonical IMF distance. Small-angle X-ray scattering and molecular dynamics simulation indicate that the interdomain linkers are tightly packed via hydrophobic interactions, which likely sustains the chirality transmission between DNA and peptide domains. Our findings establish that long-range chiral transmission occurs in tertiary macromolecular domains, explaining the presence of homochiral pairing of superhelices in proteins.

doi.org/10.1038/s41467-021-27708-4 - 2022

PubReading [150] - 1‐Deazaguanosine-Modified RNA: The Missing Piece for Functional RNA Atomic Mutagenesis - R. Bereiter, R. Micura et al

Tue, 21 Jun 2022 20:00:18 GMT

Atomic mutagenesis is the key to advance our understanding of RNA recognition and RNA catalysis. To this end, deazanucleosides are utilized to evaluate the participation of specific atoms in these processes. One of the remaining challenges is access to RNA-containing 1-deazaguanosine (c1G). Here, we present the synthesis of this nucleoside and its phosphoramidite, allowing first time access to c1G-modified RNA. Thermodynamic analyses revealed the base pairing parameters for c1G-modified RNA. Furthermore, by NMR spectroscopy, a c1G-triggered switch of Watson-Crick into Hoogsteen pairing in HIV-2 TAR RNA was identified. Additionally, using X-ray structure analysis, a guanine−phosphate backbone interaction affecting RNA fold stability was characterized, and finally, the critical impact of an active-site guanine in twister ribozyme on the phosphodiester cleavage was revealed. Taken together, our study lays the synthetic basis for c1G-modified RNA and demonstrates the power of the completed deazanucleoside toolbox for RNA atomic mutagenesis needed to achieve in-depth understanding of RNA recognition and catalysis.

doi.org/10.1021/jacs.2c01877 - 2022

PubReading [149] - Therapeutic design of peptide modulators of protein-protein interactions in membranes - T. Stone and C. Deber

Mon, 20 Jun 2022 20:00:20 GMT

Membrane proteins play the central roles in a variety of cellular processes, ranging from nutrient uptake and signalling, to cell-cell communication. Their biological functions are directly related to how they fold and assemble; defects often lead to disease. Protein–protein interactions (PPIs) within the membrane are therefore of great interest as therapeutic targets. Here we review the progress in the application of membrane–insertable peptides for the disruption or stabilization of membrane–based PPIs. We describe the design and preparation of transmembrane peptide mimics; and of several categories of peptidomimetics used for study, including D-enantiomers, non–natural amino acids, peptoids, and β-peptides. Further aspects of the review describe modifications to membrane–insertable peptides, including lipidation and cyclization via hydrocarbon stapling. These approaches provide a pathway toward the development of metabolically stable, non-toxic, and efficacious peptide modulators of membrane–based PPIs. This article is part of a Special Issue entitled: Lipid order/lipid defects and lipid- control of protein activity edited by Dirk Schneider.

doi.org/10.1016/j.bbamem.2016.08.013 - 2017

PubReading [148] - The current landscape of nucleic acid therapeutics - J. Kulkarni, R. van der Meel et al.

Fri, 17 Jun 2022 20:00:18 GMT

The increasing number of approved nucleic acid therapeutics demonstrates the potential to treat diseases by targeting their genetic blueprints in vivo. Conventional treatments generally induce therapeutic effects that are transient because they target proteins rather than underlying causes. In contrast, nucleic acid therapeutics can achieve long-lasting or even curative effects via gene inhibition, addition, replacement or editing. Their clinical translation, however, depends on delivery technologies that improve stability, facilitate internalization and increase target affinity. Here, we review four platform technologies that have enabled the clinical translation of nucleic acid therapeutics: antisense oligonucleotides, ligand-modified small interfering RNA conjugates, lipid nanoparticles and adeno-associated virus vectors. For each platform, we discuss the current state-of-the-art clinical approaches, explain the rationale behind its development, highlight technological aspects that facilitated clinical translation and provide an example of a clinically relevant genetic drug. In addition, we discuss how these technologies enable the development of cutting-edge genetic drugs, such as tissue-specific nucleic acid bioconjugates, messenger RNA and gene-editing therapeutics.

doi.org/10.1038/s41565-021-00898-0 - 2021

PubReading [147] - Furan-containing polymeric Materials: Harnessing the Diels-Alder chemistry for biomedical applications - T. Gevrek and A. Sanyal

Thu, 16 Jun 2022 20:00:18 GMT

Polymeric materials that can be readily modified with (bio)molecules of interest to render them suitable for intended applications are in demand for use in various areas of biomedical and materials science. Among various clickable reactive groups that can be incorporated into polymeric materials as handles for functionalization, ready availability, efficient reactivity under relatively mild conditions, as well as selectivity to specific functional groups makes the furan moiety an attractive choice. While the dynamic nature of furan-maleimide cycloadduct has been exploited for the fabrication of self-healing materials for decades, this reversibility has been only recently exploited to further expand the functional properties of polymeric materials. This review article focuses on recent examples that demonstrate that functional properties imparted to polymeric materials through incorporation of the furan group go beyond self-healing. Recent examples illustrating biomolecular immobilization, drug delivery/release, and dynamic material fabrication are highlighted to show the versatility of furan- containing polymeric materials in various biomedical applications.

doi.org/10.1016/j.eurpolymj.2021.110514 - 2021

PubReading [146] - Pyrene-modified PNAs: Stacking interactions and selective excimer emission in PNA2DNA triplexes - A. Manicardi, L. Guidi, A. Ghidini and R. Corradini

Wed, 15 Jun 2022 20:00:19 GMT

Pyrene derivatives can be incorporated into nucleic acid analogs in order to obtain switchable probes or supramolecular architectures. In this paper, peptide nucleic acids (PNAs) containing 1 to 3 1-pyreneacetic acid units (PNA1–6) with a sequence with prevalence of pyrimidine bases, complementary to cystic fibrosis W1282X point mutation were synthesized. These compounds showed sequence-selective switch-on of pyrene excimer emission in the presence of target DNA, due to PNA2DNA triplex formation, with stability depending on the number and positioning of the pyrene units along the chain. An increase in triplex stability and a very high mismatch-selectivity, derived from combined stacking and base-pairing interactions, were found for PNA2, bearing two distant pyrene units.

doi:10.3762/bjoc.10.154 - 2014

PubReading [145] - CAR T cell therapy and the tumor microenvironment: Current challenges and opportunities - L. Fonkoua, S. Kenderian et al

Tue, 14 Jun 2022 20:00:18 GMT

Chimeric antigen receptor (CAR) T cell therapy has demonstrated remarkable outcomes in individuals with hematological malignancies, but its success has been hindered by barriers intrinsic to the tumor microenvironment (TME), particularly for solid tumors, where it has yet to make its mark. In this article, we provide an updated review and future perspectives on features of the TME that represent barriers to CART cell therapy efficacy, including competition for metabolic fuels, physical barriers to infiltration, and immunosuppressive factors. We then discuss novel and promising strategies to overcome these obstacles that are in preclinical development or under clinical investigation.

doi.org/10.1016/j.omto.2022.03.009 - 2022

PubReading [144] - What machine learning can do for developmental biology - P. Villoutreix

Mon, 13 Jun 2022 20:00:18 GMT

Developmental biology has grown into a data intensive science with the development of high-throughput imaging and multi-omics approaches. Machine learning is a versatile set of techniques that can help make sense of these large datasets with minimal human intervention, through tasks such as image segmentation, super- resolution microscopy and cell clustering. In this Spotlight, I introduce the key concepts, advantages and limitations of machine learning, and discuss how these methods are being applied to problems in developmental biology. Specifically, I focus on how machine learning is improving microscopy and single-cell ‘omics’ techniques and data analysis. Finally, I provide an outlook for the futures of these fields and suggest ways to foster new interdisciplinary developments.

oi:10.1242/dev.188474 - 2021

PubReading [143] - Homology directed correction, a new pathway model for point mutation repair catalyzed by CRISPR‐Cas - B. Sansbury, E. Kmiec et al

Fri, 10 Jun 2022 20:00:18 GMT

Gene correction is often referred to as the gold standard for precise gene editing and while CRISPR‐ Cas systems continue to expand the toolbox for clinically relevant genetic repair, mechanistic hurdles still hinder widespread implementation. One of the most prominent challenges to precise CRISPR‐ directed point mutation repair centers on the prevalence of on‐site mutagenesis, wherein insertions and deletions appear at the targeted site following correction. Here, we introduce a pathway model for Homology Directed Correction, specifically point mutation repair, which enables a foundational analysis of genetic tools and factors influencing precise gene editing. To do this, we modified an in vitro gene editing system which utilizes a cell‐free extract, CRISPR‐Cas RNP and donor DNA template to catalyze point mutation repair. We successfully direct correction of four unique point mutations which include two unique nucleotide mutations at two separate targeted sites and visualize the repair profiles resulting from these reactions. This extension of the cell‐free gene editing system to model point mutation repair may provide insight for understanding the factors influencing precise point mutation correction.

doi.org/10.1038/s41598-022-11808-2 - 2022

PubReading [142] - Artificial nucleic acid backbones and their applications in therapeutics, synthetic biology and biotechnology - S. Epple, A. El-Sagheer and T. Brown

Thu, 09 Jun 2022 20:00:19 GMT

The modification of DNA or RNA backbones is an emerging technology for therapeutic oligonucleotides, synthetic biology and biotechnology. Despite a plethora of reported arti- ficial backbones, their vast potential is not fully utilised. Limited synthetic accessibility remains a major bottleneck for the wider application of backbone-modified oligonucleo- tides. Thus, a variety of readily accessible artificial backbones and robust methods for their introduction into oligonucleotides are urgently needed to utilise their full potential in therapeutics, synthetic biology and biotechnology.

doi.org/10.1042/ETLS20210169 - 2021

PubReading [141] - Single-molecule fluorescence detection of a tricyclic nucleoside analogue - G. Samaan, B. Purse et al

Wed, 08 Jun 2022 20:00:18 GMT

Fluorescent nucleobase surrogates capable of Watson–Crick hydrogen bonding are essential probes of nucleic acid structure and dynamics, but their limited brightness and short absorption and emission wavelengths have rendered them unsuitable for single-molecule detection. Aiming to improve on these properties, we designed a new tricyclic pyrimidine nucleoside analogue with a push–pull conjugated system and synthesized it in seven sequential steps. The resulting C-linked 8-(diethylamino)benzo[b][1,8] naphthyridin-2(1H)-one nucleoside, which we name ABN, exhibits 3442 1⁄4 20 000 M1 cm1 and Fem,540 1⁄4 0.39 in water, increasing to Fem 1⁄4 0.50–0.53 when base paired with adenine in duplex DNA oligonucleotides. Single-molecule fluorescence measurements of ABN using both one-photon and two- photon excitation demonstrate its excellent photostability and indicate that the nucleoside is present to > 95% in a bright state with count rates of at least 15 kHz per molecule. This new fluorescent nucleobase analogue, which, in duplex DNA, is the brightest and most red-shifted known, is the first to offer robust and accessible single-molecule fluorescence detection capabilities.

DOI: 10.1039/d0sc03903a - 2021

PubReading [140] - Peptide Nucleic Acid-Based Biosensors for Cancer Diagnosis - R. D’Agata, M. Giuffrida and G. Spoto

Tue, 07 Jun 2022 20:00:18 GMT

The monitoring of DNA and RNA biomarkers freely circulating in the blood constitutes the basis of innovative cancer detection methods based on liquid biopsy. Such methods are expected to provide new opportunities for a better understanding of cancer disease at the molecular level, thus contributing to improved patient outcomes. Advanced biosensors can advance possibilities for cancer-related nucleic acid biomarkers detection. In this context, peptide nucleic acids (PNAs) play an important role in the fabrication of highly sensitive biosensors. This review provides an overview of recently described PNA-based biosensors for cancer biomarker detection. One of the most striking features of the described detection approaches is represented by the possibility to detect target nucleic acids at the ultra-low concentration with the capability to identify single-base mutations.

doi: 10.3390/molecules22111951 - 2017

PubReading [139] - Harnessing DNA as a Designable Scaffold for Asymmetric Catalysis: Recent Advances and Future Perspectives - JJ. Yum, H. Sugiyama and S. Park

Mon, 06 Jun 2022 20:00:18 GMT

Since the first report of DNAzyme by in vitro selection in 1994, catalytic DNA has investigated extensively, and their application has expanded continually in virtue of rapid advances in molecular biology and biotechnology. Nowadays, DNA is in the second prime time by way of DNA-based hybrid catalysts and DNA metalloenzymes in which helical chirality of DNA serves to asymmetric catalysis. DNA-based hybrid catalysts are attractive system to respond the demand of the times to pursuit green and sustainable society beyond traditional catalytic systems that value reaction efficiency. Herein, we highlight the recent advances and perspective of DNA-based hybrid catalysts with various aspects of DNA as a versatile scaffold for asymmetric synthesis. We hope that scientists in a variety of fields will be encouraged to join and promote remarkable evolution of this interesting research.

doi.org/10.1002/tcr.202100333 - 2022

PubReading [138] - Improving CRISPR–Cas specificity with chemical modifications in single-guide RNAs - D. Ryan, D. Dellinger et al

Fri, 03 Jun 2022 20:00:19 GMT

CRISPR systems have emerged as transformative tools for altering genomes in living cells with un- precedented ease, inspiring keen interest in increasing their specificity for perfectly matched targets. We have developed a novel approach for improving specificity by incorporating chemical modifications in guide RNAs (gRNAs) at specific sites in their DNA recognition sequence (‘guide sequence’) and systematically evaluating their on-target and off-target activities in biochemical DNA cleavage assays and cell-based assays. Our results show that a chemical modification (2′-O-methyl-3′-phosphonoacetate, or ‘MP’) incorporated at select sites in the ribose- phosphate backbone of gRNAs can dramatically re- duce off-target cleavage activities while maintaining high on-target performance, as demonstrated in clinically relevant genes. These findings reveal a unique method for enhancing specificity by chemically modifying the guide sequence in gRNAs. Our approach introduces a versatile tool for augmenting the performance of CRISPR systems for research, industrial and therapeutic applications.

doi: 10.1093/nar/gkx1199 - 2018

PubReading [137] - Double-headed nucleic acids condense the molecular information of DNA to half the number of nucleotides -K. Beck, P. Nielsen et al

Thu, 02 Jun 2022 20:00:18 GMT

Nucleotide monomers that hold two nucleobases each, i.e. double- headed nucleotides, have been shown to form two sets of functional Watson–Crick base pairs when incorporated into dsDNA, and they hereby behave as dinucleotides. To form the basis for fully modified double-headed nucleic acids (DhNA), we have prepared three new DhNA monomers and can now demonstrate that the molecular information of 10 Watson–Crick base pairs can be condensed to highly stable 5-mer DhNA duplexes.

DOI: 10.1039/d1cc03539h - 2021

PubReading [136] - A prebiotically plausible scenario of an RNA–peptide world - F. Müller, C. Chan, T. Carell

Wed, 01 Jun 2022 20:00:18 GMT

The RNA world concept1 is one of the most fundamental pillars of the origin of life theory. It predicts that life evolved from increasingly complex self-replicating RNA molecules. The question of how this RNA world then advanced to the next stage, in which proteins became the catalysts of life and RNA reduced its function predominantly to information storage, is one of the most mysterious chicken-and-egg conundrums in evolution. Here we show that non-canonical RNA bases, which are found today in transfer and ribosomal RNAs, and which are considered to be relics of the RNA world, are able to establish peptide synthesis directly on RNA. The discovered chemistry creates complex peptide-decorated RNA chimeric molecules, which suggests the early existence of an RNA–peptide world13 from which ribosomal peptide synthesis14 may have emerged. The ability to grow peptides on RNA with the help of non-canonical vestige nucleosides offers the possibility of an early co-evolution of covalently connected RNAs and peptides, which then could have dissociated at a higher level of sophistication to create the dualistic nucleic acid–protein world that is the hallmark of all life on Earth.

doi.org/10.1038/s41586-022-04676-3 - 2022

PubReading [135] - Structural basis for broad anti-phage immunity by DISARM - J. Bravo, D. Taylor et al

Tue, 31 May 2022 20:00:18 GMT

In the evolutionary arms race against phage, bacteria have assembled a diverse arsenal of antiviral immune strategies. While the recently discovered DISARM (Defense Island System Associated with Restriction-Modification) systems can provide protection against a wide range of phage, the molecular mechanisms that underpin broad antiviral targeting but avoiding autoimmunity remain enigmatic. Here, we report cryo-EM structures of the core DISARM complex, DrmAB, both alone and in complex with an unmethylated phage DNA mimetic. These structures reveal that DrmAB core complex is autoinhibited by a trigger loop (TL) within DrmA and binding to DNA substrates containing a 5′ overhang dislodges the TL, initiating a long-range structural rearrangement for DrmAB activation. Together with structure-guided in vivo studies, our work provides insights into the mechanism of phage DNA recognition and specific activation of this widespread antiviral defense system.

doi.org/10.1038/s41467-022-30673-1 - 2022

PubReading [134] - Tethering Carbohydrates to the Vinyliminium Ligand of Antiproliferative Organometallic Diiron Complexes - S. Schoch, M. Dalla Pozza, G. Gasser, F. Marchetti et al

Mon, 30 May 2022 20:00:17 GMT

Four propargyl O-glycosides derivatized with mannose, glucose, and fructose moieties were synthesized and then incorporated within a diiron structure as part of a vinyliminium ligand. Hence, six glycoconjugated diiron complexes, [2−5]CF3SO3 (see Scheme 1) and the nonglycosylated analogues [6a−b]CF3SO3, were obtained in high yields and unambiguously characterized by elemental analysis, mass spectrometry, and IR and multinuclear NMR spectroscopies. All compounds exhibited a significant stability in DMSO-d6/D2O solution, with 63−89% of the complexes unaltered after 72 h at 37 °C and also in the cell culture medium. The cytotoxicity of [2−6]CF3SO3, as well as that of previously reported 7 and 8, was assessed on CT26 (mouse colon carcinoma), U87 (human glioblastoma), MCF-7 (human breast adenocarcinoma), and RPE-1 (human normal retina pigmented epithelium) cell lines. In general, the IC50 values correlate with the hydrophobicity of the compounds (measured as octanol−water partition coefficients) and do not show an appreciable level of selectivity against cancer cells with respect to the nontumor ones.

doi.org/10.1021/acs.organomet.1c00519 - 2022

PubReading [133] - Life in the light: nucleic acid photoproperties as a legacy of chemical evolution - A. Beckstead, B. Kohler

Thu, 26 May 2022 20:00:18 GMT

Photophysical investigations of the canonical nucleobases that make up DNA and RNA during the past 15 years have revealed that excited states formed by the absorption of UV radiation decay with subpicosecond lifetimes (i.e., o10􏰁12 s). Ultrashort lifetimes are a general property of absorbing sunscreen molecules, suggesting that the nucleobases are molecular survivors of a harsh UV environment. Encoding the genome using photostable building blocks is an elegant solution to the threat of photochemical damage. Ultrafast excited-state deactivation strongly supports the hypothesis that UV radiation played a major role in shaping molecular inventories on the early Earth before the emergence of life and the subsequent development of a protective ozone shield. Here, we review the general physical and chemical principles that underlie the photostability, or ‘‘UV hardiness’’, of modern nucleic acids and discuss the possible implications of these findings for prebiotic chemical evolution. In RNA and DNA strands, much longer-lived excited states are observed, which at first glance appear to increase the risk of photo-chemistry. It is proposed that the dramatically different photoproperties that emerge from assemblies of photostable building blocks may explain the transition from a world of molecular survival to a world in which energy-rich excited electronic states were eventually tamed for biological purposes such as energy transduction, signaling, and repair of the genetic machinery. - DOI: 10.1039/c6cp04230a - 2016

PubReading [132] - Unnatural bases for recognition of noncoding nucleic acid interfaces - S. Miao, D. Bong et al

Wed, 25 May 2022 20:00:18 GMT

The notion of using synthetic heterocycles instead of the native bases to interface with DNA and RNA has been explored for nearly 60 years. Unnatural bases compatible with the DNA/RNA coding interface have the potential to expand the genetic code and co-opt the machinery of biology to access new macromolecular function; accordingly, this body of research is core to synthetic biology. While much of the literature on artificial bases focuses on code expansion, there is a significant and grow- ing effort on docking synthetic heterocycles to noncoding nucleic acid interfaces; this approach seeks to illuminate major processes of nucleic acids, including regulation of transcription, translation, transport, and transcript lifetimes. These major avenues of research at the coding and noncoding interfaces have in common fundamental principles in molecular recognition. Herein, we provide an overview of foundational literature in biophysics of base recognition and unnatural bases in coding to provide context for the developing area of targeting noncoding nucleic acid interfaces with synthetic bases, with a focus on systems developed through iterative design and biophysical study.

PubReading [131] - Microneedling for Hair Loss - A. Gupta, M. Bamimore et al.

Tue, 24 May 2022 20:00:18 GMT

There are limited studies that investigate microneedling as a monotherapy for hair loss since majority of the trials combine it with other therapies such as topical minoxidil or platelet-rich plasma. While preliminary results look promising, further investigation of microneedling as a monotherapy in larger, randomized controlled trials will help determine its safety and efficacy, and place in treating Androgenic Alopecia. - DOI: 10.1111/jocd.14525 - 2021

PubReading [130] - Computational analysis of next generation sequencing data and its applications in clinical oncology - R. Wadapurkar and Renu Vyas

Mon, 23 May 2022 20:00:19 GMT

Next generation sequencing (NGS) has made great strides in sequencing technology as it enables sequencing of genes in a high throughput manner with low cost. Various NGS platforms such as Illumina, Roche, ABI/SOLiD are used for wet-lab analysis of NGS data and computational tools such as BWA, Bowtie, Galaxy, SanGeniX are used for dry-lab analysis of NGS data. One of the important aspects of NGS data is its usage in early disease diagnosis especially in cancer which was earlier not possible with conventional sequencing technologies such as Sanger sequencing, NGS can identify all those mutations which cannot be identified using conventional sequencing technologies as researchers can now sequence the whole genome, exome or transcriptome. Exome sequencing is preferred, as a higher number of mutations are found to exist in the exome part of genes. The present comprehensive review encompasses the complete NGS data analysis workflow that includes alignment of NGS reads, identification and annotation of mutations and visualization, discussion of software tools for variant identification and annotation, evaluation of structural variation in NGS data, and study of different DNA sequencing technologies. In the field of clinical oncology, NGS has already proven its usefulness, and the mortality rate has been reduced, as now doctors can suggest a proper treatment to their patients by checking the complete genomic profile. However, data storage and the complexity in interpreting enormous amounts of data obtained with NGS still remain a computational challenge to researchers, as for each sample, the number of different and very large analysis files are generated directly from the raw sequence read file to the final result file. NGS resultant data is very complex, and its interpretation requires expert bioinformatics assistance, as a large number of mutations are identified from samples, but to differentiate clinically significant mutations among them with appropriate use of validation methods is a challenging task. This review is intended to provide researchers with a complete overview of NGS along with knowledge of how the tools will be employed, and insight into identification and interpretation of cancer mutations for clinical diagnostics. - doi.org/10.1016/j.imu.2018.05.003 - 2018

PubReading [129] - Beginner’s guide to mass spectrometry–based proteomics - A. Sinha and M. Mann

Fri, 20 May 2022 20:00:18 GMT

Mass spectrometry (MS)-based proteomics is the most comprehensive approach for the quantitative profiling of proteins, their interactions and modifications. It is a challenging topic as a firm grasp requires expertise in biochemistry for sample preparation, analytical chemistry for instrumentation and computational biology for data analysis. In this short guide, we highlight the various components of a mass spectrometer, the sample preparation process for conversion of proteins into peptides, and quantification and analysis strategies. The advancing technology of MS-based proteomics now opens up opportunities in clinical applications and single-cell analysis. - doi.org/10.1042/BIO20200057 - 2020

PubReading [128] - Best practices for variant calling in clinical sequencing - D. Koboldt

Thu, 19 May 2022 20:00:19 GMT

Next-generation sequencing technologies have enabled a dramatic expansion of clinical genetic testing both for inherited conditions and diseases such as cancer. Accurate variant calling in NGS data is a critical step upon which virtually all downstream analysis and interpretation processes rely. Just as NGS technologies have evolved considerably over the past 10 years, so too have the software tools and approaches for detecting sequence variants in clinical samples. In this review, I discuss the current best practices for variant calling in clinical sequencing studies, with a particular emphasis on trio sequencing for inherited disorders and somatic mutation detection in cancer patients. I describe the relative strengths and weaknesses of panel, exome, and whole-genome sequencing for variant detection. Recommended tools and strategies for calling variants of different classes are also provided, along with guidance on variant review, validation, and benchmarking to ensure optimal performance. Although NGS technologies are continually evolving, and new capabilities (such as long-read single-molecule sequencing) are emerging, the “best practice” principles in this review should be relevant to clinical variant calling in the long term. - doi.org/10.1186/s13073-020-00791-w - 2020

PubReading [188] - m1A and m6A modifications function cooperatively to facilitate rapid mRNA degradation - S. Ho Boo, H. Ha, Y. Kim

Wed, 28 Sep 2022 20:00:17 GMT

N6-Methyladenosine (m6A), the most abundant internal mRNA modification, affects multiple steps in gene expression. Mechanistically, the binding of YTHDF2 to m6A on mRNAs elicits rapid mRNA degradation by recruiting several RNA degrading enzymes. Here, we show that N1-methyladenosine (m1A), another type of RNA modification, accelerates rapid m6A RNA degradation. We identify HRSP12 as an RNA-binding protein that recognizes m1A. The binding of HRSP12 to m1A promotes efficient interaction of YTHDF2 with m6A, consequently facilitating endoribonucleolytic cleavage via the RNase P/MRP complex. Transcriptome-wide analyses also reveal that mRNAs harboring both m1A and m6A are downregulated in an HRSP12-dependent manner compared with mRNAs harboring m6A only. Accordingly, a subset of endogenous circular RNAs that harbor m6A and associate with YTHDF2 in an HRSP12-dependent manner is also subjected to m1A-facilitated rapid degradation. Together, our observations provide compelling evidence for crosstalk between different RNA modifications.

doi.org/10.1016/j.celrep.2022.111317 - 2022

PubReading [127] - Insufficient Evidence for ‘‘Autism-Specific’’ Genes - S. Myers, D. Ledbetter

Wed, 18 May 2022 20:00:18 GMT

Despite evidence that deleterious variants in the same genes are implicated across multiple neurodevelopmental and neuropsychiatric disorders, there has been considerable interest in identifying genes that, when mutated, confer risk that is largely specific for autism spectrum disorder (ASD). Here, we review the findings and limitations of recent efforts to identify relatively ‘‘autism-specific’’ genes, efforts which focus on rare variants of large effect size that are thought to account for the observed phenotypes. We present a divergent interpretation of published evidence; discuss practical and theoretical issues related to studying the relationships between rare, large-effect deleterious variants and neurodevelopmental phenotypes; and describe potential future directions of this research. We argue that there is currently insufficient evidence to establish meaningful ASD specificity of any genes based on large-effect rare-variant data. - doi.org/10.1016/j.ajhg.2020.04.004 - 2020

PubReading [126] - Split light up aptamers as a probing tool for nucleic acids - Y. Gerasimovaa, D Nedorezovab and D. Kolpashchikov

Tue, 17 May 2022 20:00:17 GMT

Aptamers that bind non-fluorescent dyes and increase their fluorescence can be converted to fluorescent sensors. Here, we discuss and provide guidance for the design of split (binary) light up aptameric sensors (SLAS) for nucleic acid analysis. SLAS consist of two RNA or DNA strands and a fluorogenic organic dye added as a buffer component. The two strands hybridize to the analyzed DNA or RNA sequence and form a dye-binding pocket, followed by dye binding, and increase in its fluorescence. SLAS can detect nucleic acids in a cost-efficient label-free format since it does not require conjugation of organic dyes with nucleic acids. SLAS design is preferable over monolith fluorescent sensors due to simpler assay optimization and improved selectivity. RNA-based SLAS can be expressed in cells and used for intracellular monitoring and imaging biological molecules.

PubReading [125] - How cryo-electron microscopy and X-ray crystallography complement each other - H. Wang, and J. Wang

Mon, 16 May 2022 20:00:19 GMT

With the ability to resolve structures of macromolecules at atomic resolution, X-ray crystallography has been the most powerful tool in modern structural biology. At the same time, recent technical improvements have triggered a resolution revolution in the single-particle cryo-EM method. While the two methods are different in many respects, from sample preparation to structure determination, they both have the power to solve macromolecular structures at atomic resolution. It is important to understand the unique advantages and caveats of the two methods in solving structures and to appreciate the complementary nature of the two methods in structural biology. In this review we provide some examples, and discuss how X-ray crystallography and cryo-EM can be combined in deciphering structures of macromolecules for our full understanding of their biological mechanisms.- DOI: 10.1002/pro.3022 - 2016

PubReading [124] - A biomedical open knowledge network harnesses the power of AI to understand deep human biology - S. Baranzini, S. Huang et al

Fri, 13 May 2022 20:00:17 GMT

Knowledge representation and reasoning (KR&R) has been successfully imple- mented in many fields to enable computers to solve complex problems with AI methods. However, its application to biomedicine has been lagging in part due to the daunting complexity of molecular and cellular pathways that govern human physiology and pathology. In this article, we describe concrete uses of Scalable PrecisiOn Medicine Knowledge Engine (SPOKE), an open knowledge network that connects curated information from thirty-seven specialized and human- curated databases into a single property graph, with 3 million nodes and 15 mil- lion edges to date. Applications discussed in this article include drug discovery, COVID-19 research and chronic disease diagnosis, and management. - DOI: 10.1002/aaai.12037 - 2021

PubReading [123] - Next-Generation Sequencing Technologies - R. McCombie, J. McPherson, and E. Mardis

Thu, 12 May 2022 20:00:17 GMT

Although DNA and RNA sequencing has a history spanning five decades, large-scale massively parallel sequencing, or next-generation sequencing (NGS), has only been commercially available for about 10 years. Nonetheless, the meteoric increase in sequencing throughput with NGS has dramatically changed our understanding of our genome and our- selves. Sequencing the first human genome as a haploid reference took nearly 10 years but now a full diploid human genome sequence can be accomplished in just a few days. NGS has also reduced the cost of generating sequence data and a plethora of sequence-based methods for probing a genome have emerged using NGS as the readout and have been applied to many species. NGS methods have also entered the medical realm and will see increasing use in diagnosis and treatment. NGS has largely been driven by short-read generation (150 bp) but new platforms have emerged and are now capable of generating long multikilobase reads. These latter platforms enable reference-independent genome assemblies and long-range haplotype generation. Rapid DNA and RNA sequencing is now mainstream and will continue to have an increasing impact on biology and medicine. - oi: 10.1101/cshperspect.a036798 - 2019

PubReading [122] - Systematic benchmarking of tools for CpG methylation detection from nanopore sequencing - Z. Yuen, E. Eyras et al

Wed, 11 May 2022 20:00:16 GMT

DNA methylation plays a fundamental role in the control of gene expression and genome integrity. Although there are multiple tools that enable its detection from Nanopore sequencing, their accuracy remains largely unknown. Here, we present a systematic benchmarking of tools for the detection of CpG methylation from Nanopore sequencing using individual reads, control mixtures of methylated and unmethylated reads, and bisulfite sequencing. We found that tools have a tradeoff between false positives and false negatives and present a high dispersion with respect to the expected methylation frequency values. We described various strategies to improve the accuracy of these tools, including a consensus approach, METEORE (https://github.com/comprna/METEORE), based on the combination of the predictions from two or more tools that shows improved accuracy over individual tools. Snakemake pipelines are also provided for reproducibility and to enable the systematic application of our analyses to other datasets. - doi.org/10.1038/s41467-021-23778-6 - 2021

PubReading [117] - Real-time Voltammetric Anion Sensing Under Flow - S. Patrick, J. Davis

Wed, 04 May 2022 20:00:17 GMT

The development of real-life applicable ion sensors, in particular those capable of repeat use and long-term monitoring, remains a formidable challenge. Herein, we demonstrate, in a proof-of-concept, the real-time voltammet- ric sensing of anions under continuous flow in a 3D-printed microfluidic system. Electro-active anion receptive halogen bonding (XB) and hydrogen bonding (HB) ferrocene-isophtha- lamide-(iodo)triazole films were employed as exemplary sensory interfaces. Upon exposure to anions, the cathodic perturbations of the ferrocene redox-transducer are monitored by repeat square-wave voltammetry (SWV) cycling and peak fitting of the voltammograms by a custom-written MATLAB script. This enables the facile and automated data processing of thousands of SW scans and is associated with an over one order-of-magnitude improvement in limits of detection. In addition, this improved analysis enables tuning of the measurement parameters such that high temporal resolution can be achieved. More generally, this new flow methodology is extendable to a variety of other analytes, including cations, and presents an important step towards translation of voltammetric ion sensors from laboratory to real-world applications. - doi.org/10.1002/chem.202103249 - 2021

PubReading [121] - Cone-shaped HIV-1 capsids are transported through intact nuclear pores - V. Zila, M. Beck et al

Tue, 10 May 2022 20:00:17 GMT

Human immunodeficiency virus (HIV-1) remains a major health threat. Viral capsid uncoating and nuclear import of the viral genome are critical for productive infection. The size of the HIV-1 capsid is generally believed to exceed the diameter of the nuclear pore complex (NPC), indicating that capsid uncoating has to occur prior to nuclear import. Here, we combined correlative light and electron microscopy with subtomogram averaging to capture the structural status of reverse transcription-competent HIV-1 complexes in infected T cells. We demonstrated that the diameter of the NPC in cellulo is sufficient for the import of apparently intact, cone-shaped capsids. Subsequent to nuclear import, we detected disrupted and empty capsid fragments, indicating that uncoating of the replication complex occurs by breaking the capsid open, and not by disassembly into individual subunits. Our data directly visualize a key step in HIV-1 replication and enhance our mechanistic understanding of the viral life cycle. - doi.org/10.1016/j.cell.2021.01.025 - 2021

PubReading [120] - Modern epigenetics methods in biological research - Yuanyuan Li

Mon, 09 May 2022 20:00:16 GMT

The definition of epigenetics refers that molecular modifications on DNA that can regulate gene activity are independent of DNA sequence and mitotically stable. Notably, epigenetics studies have grown exponentially in the past few years. Recent progresses that lead to exciting discoveries and groundbreaking nature of this area demand thorough methodologies and advanced technologies to move epigenetics to the forefront of molecular biology. The most recognized epigenetic regulations are DNA methylation, histone modifications, and non-coding RNAs (ncRNAs). This review will discuss the modern techniques that are available to detect locus-specific and genome-wide changes for all epigenetic codes. Furthermore, updated analysis of technologies, newly developed methods, recent breakthroughs and bioinformatics pipelines in epigenetic analysis will be presented. These methods, as well as many others presented in this specific issue, provide comprehensive guidelines in the area of epigenetics that facilitate further developments in this promising and rapidly developing field. - doi.org/10.1016/j.ymeth.2020.06.022 - 2020

PubReading [119] - CAR T‐cell therapy: Full speed ahead - D. Sermer and R. Brentjens

Fri, 06 May 2022 20:00:16 GMT

Chimeric antigen receptor (CAR) T‐cell therapy has dramatically shifted the landscape of treatment for lymphoid malignancies, especially diffuse large B‐cell lymphoma (DLBCL) and acute lymphoblastic leukemia (ALL). However, there continue to be significant limitations of this therapy, such as incomplete or nonsustained responses and severe toxicities in a subset of patients. Furthermore, expanding the role of CAR T‐cell therapy to new disease types is an important next step. In this review, we will highlight landmark trials for anti‐CD19 CAR T cells and first‐in‐human trials of novel CARs, as well as discuss promising innovative CAR designs that are still undergoing preclinical development. Lastly, we will discuss toxicity and mechanisms of CAR T‐cell resistance and failure, as well as potential future treatment approaches to these common issues. - DOI: 10.1002/hon.2591 - 2019

PubReading [118] - Robust transcriptome-wide discovery of RNA-binding protein binding sites with enhanced CLIP (eCLIP) - E. Van Nostrand, G. Yeo et al

Thu, 05 May 2022 20:00:16 GMT

As RNA-binding proteins (RBPs) play essential roles in cellular physiology by interacting with target RNA molecules, binding site identification by UV crosslinking and immunoprecipitation (CLIP) of ribonucleoprotein complexes is critical to understanding RBP function. However, current CLIP protocols are technically demanding and yield low-complexity libraries with high experimental failure rates. We have developed an enhanced CLIP (eCLIP) protocol that decreases requisite amplification by ∼1,000-fold, decreasing discarded PCR duplicate reads by ∼60% while maintaining single-nucleotide binding resolution. By simplifying the generation of paired IgG and size-matched input controls, eCLIP improves specificity in the discovery of authentic binding sites. We generated 102 eCLIP experiments for 73 diverse RBPs in HepG2 and K562 cells (available at https://www.encodeproject.org), demonstrating that eCLIP enables large-scale and robust profiling, with amplification and sample requirements similar to those of ChIP-seq. eCLIP enables integrative analysis of diverse RBPs to reveal factor-specific profiles, common artifacts for CLIP and RNA-centric perspectives on RBP activity. - DOI: 10.1038/nmeth.3810 - 2016

PubReading [116] - Improved protein structure prediction using potentials from deep learning - A. Senior, D. Hassabis et al.

Tue, 03 May 2022 20:00:19 GMT

Protein structure prediction can be used to determine the three-dimensional shape of a protein from its amino acid sequence1. This problem is of fundamental importance as the structure of a protein largely determines its function2; however, protein structures can be difficult to determine experimentally. Considerable progress has recently been made by leveraging genetic information. It is possible to infer which amino acid residues are in contact by analysing covariation in homologous sequences, which aids in the prediction of protein structures3. Here we show that we can train a neural network to make accurate predictions of the distances between pairs of residues, which convey more information about the structure than contact predictions. Using this information, we construct a potential of mean force4 that can accurately describe the shape of a protein. We find that the resulting potential can be optimized by a simple gradient descent algorithm to generate structures without complex sampling procedures. The resulting system, named AlphaFold, achieves high accuracy, even for sequences with fewer homologous sequences. In the recent Critical Assessment of Protein Structure Prediction5 (CASP13)—a blind assessment of the state of the field—AlphaFold created high-accuracy structures (with template modelling (TM) scores6 of 0.7 or higher) for 24 out of 43 free modelling domains, whereas the next best method, which used sampling and contact information, achieved such accuracy for only 14 out of 43 domains. AlphaFold represents a considerable advance in protein-structure prediction. We expect this increased accuracy to enable insights into the function and malfunction of proteins, especially in cases for which no structures for homologous proteins have been experimentally determined. - doi.org/10.1038/s41586-019-1923-7 - 2020

PubReading [115] - The Current Revolution in Cryo-EM - E. Egelman

Mon, 02 May 2022 20:00:16 GMT

Structural biology is the study of the molecular architecture of proteins and nucleic acids, which are the basis for all life forms. Knowledge of these structures alone is not enough to understand their functions, but it has become clear that a detailed mechanistic picture of function is not possible without structural information. Studying structure can reveal how molecules have evolved, and this type of insight would otherwise be lost by looking at only the molecule’s sequence. - .doi.org/10.1016/j.bpj.2016.02.001 - 2016

PubReading [114] - Oncogene-induced DNA damage: cyclic AMP steps into the ring - J. Fagin and J. Petrini

Fri, 29 Apr 2022 20:00:17 GMT

Growth hormone–secreting (GH-secreting) pituitary tumors are driven by oncogenes that induce cAMP signaling. In this issue of the JCI, Ben-Shlomo et al. performed a whole-exome study of pituitary adenomas. GH-secreting tumors had a high frequency of whole chromosome or chromosome arm copy number alterations and were associated with an increase in the tumor protein p53 and the cyclin-dependent kinase inhibitor p21WAF1/CIP1, which are findings consistent with induction of a response to DNA damage. Further, treatment of mouse pituitary cells with cAMP pathway agonists in vitro and in vivo elicited biomarkers of DNA replication stress or double-strand breaks. The findings of Ben Shlomo et al. indicate that oncoproteins that drive constitutively high cAMP signaling pathway output in susceptible cell types can elicit DNA replication stress and may promote genomic instability. - doi.org/10.1172/JCI142237. - 2020

PubReading [112] - Automated Tools to Advance High-Resolution Imaging in Liquid - G. Jonaid, D. Kelly

Wed, 27 Apr 2022 20:00:17 GMT

Liquid-electron microscopy (EM), the room-temperature correlate to cryo-EM, is a rapidly growing field providing high-resolution insights of macromolecules in solution. Here, we describe how liquid-EM experiments can incorporate automated tools to propel the field to new heights. We demonstrate fresh workflows for specimen preparation, data collection, and computing processes to assess biological structures in liquid. Adeno-associated virus (AAV) and the SARS-CoV-2 nucleocapsid (N) were used as model systems to highlight the technical advances. These complexes were selected based on their major differences in size and natural symmetry. AAV is a highly symmetric, icosahedral assembly with a particle diameter of ∼25 nm. At the other end of the spectrum, N protein is an asymmetric monomer or dimer with dimensions of approximately 5–7 nm, depending upon its oligomerization state. Equally important, both AAV and N protein are popular subjects in biomedical research due to their high value in vaccine development and therapeutic efforts against COVID-19. Overall, we demonstrate how automated practices in liquid-EM can be used to decode molecules of interest for human health and disease. - doi:10.1017/S1431927621013921 - 2022

PubReading [113] - Highly accurate protein structure prediction with AlphaFold - J. Jumper, P. Kohli, D. Hassabis et al

Thu, 28 Apr 2022 20:00:18 GMT

Proteins are essential to life, and understanding their structure can facilitate a mechanistic understanding of their function. Through an enormous experimental effort, the structures of around 100,000 unique proteins have been determined5, but this represents a small fraction of the billions of known protein sequences. Structural coverage is bottlenecked by the months to years of painstaking effort required to determine a single protein structure. Accurate computational approaches are needed to address this gap and to enable large-scale structural bioinformatics. Predicting the three-dimensional structure that a protein will adopt based solely on its amino acid sequence—the structure prediction component of the ‘protein folding problem’—has been an important open research problem for more than 50 years9. Despite recent progress existing methods fall far short of atomic accuracy, especially when no homologous structure is available. Here we provide the first computational method that can regularly predict protein structures with atomic accuracy even in cases in which no similar structure is known. We validated an entirely redesigned version of our neural network-based model, AlphaFold, in the challenging 14th Critical Assessment of protein Structure Prediction (CASP14), demonstrating accuracy competitive with experimental structures in a majority of cases and greatly outperforming other methods. Underpinning the latest version of AlphaFold is a novel machine learning approach that incorporates physical and biological knowledge about protein structure, leveraging multi-sequence alignments, into the design of the deep learning algorithm. - doi.org/10.1038/s41586-021-03819-2 - 2021

PubReading [111] - Chemical reprogramming of human somatic cells to pluripotent stem cells - J. Guang, H. Dang et al

Tue, 26 Apr 2022 20:00:18 GMT

Cellular reprogramming can manipulate the identity of cells to generate the desired cell types1–3. The use of cell intrinsic components, including oocyte cytoplasm and transcription factors, can enforce somatic cell reprogramming to pluripotent stem cells4–7. By contrast, chemical stimulation by exposure to small molecules offers an alternative approach that can manipulate cell fate in a simple and highly controllable manner8–10. However, human somatic cells are refractory to chemical stimulation owing to their stable epigenome2,11,12 and reduced plasticity13,14; it is therefore challenging to induce human pluripotent stem cells by chemical reprogramming. Here we demonstrate, by creating an intermediate plastic state, the chemical reprogramming of human somatic cells to human chemically induced pluripotent stem cells that exhibit key features of embryonic stem cells. The whole chemical reprogramming trajectory analysis delineated the induction of the intermediate plastic state at the early stage, during which chemical-induced dedifferentiation occurred, and this process was similar to the dedifferentiation process that occurs in axolotl limb regeneration. Moreover, we identified the JNK pathway as a major barrier to chemical reprogramming, the inhibition of which was indispensable for inducing cell plasticity and a regeneration-like program by suppressing pro-inflammatory pathways. Our chemical approach provides a platform for the generation and application of human pluripotent stem cells in biomedicine. This study lays foundations for developing regenerative therapeutic strategies that use well-defined chemicals to change cell fates in humans. - doi.org/10.1038/s41586-022-04593-5 - 2022

PubReading [110] - Hallmarks of Cancer: New Dimensions - Douglas Hanahan

Mon, 25 Apr 2022 20:00:20 GMT

The hallmarks of cancer conceptualization is a heuristic tool for distilling the vast complexity of cancer phenotypes and genotypes into a provisional set of underlying principles. As knowledge of cancer mechanisms has progressed, other facets of the disease have emerged as potential refinements. Herein, the prospect is raised that phenotypic plasticity and disrupted differentiation is a discrete hallmark capability, and that nonmutational epigenetic reprogram- ming and polymorphic microbiomes both constitute distinctive enabling characteristics that facilitate the acquisition of hallmark capabilities. Additionally, senescent cells, of varying origins, may be added to the roster of functionally important cell types in the tumor microenvironment. - doi: 10.1158/2159-8290.CD-21-1059 - 2022

PubReading [109] - Novel therapeutic targets for cancer metastasis - K. Stoletov, P. Beatty and J. Lewis

Fri, 22 Apr 2022 20:00:21 GMT

The development of metastatic disease is a complex interplay of genetic and epigenetic factors from the host and cancer cells acting in a patient-specific manner. Inhibiting key driver traits of metastasis should yield survival benefit at any stage of the disease, and we look forward to the next generation of personalized medicines for cancer therapy that target cancer cell motility for increased therapeutic efficacy. - doi.org/10.1080/14737140.2020.1718496 - 2020

PubReading [108] - Untapped Neuroimaging Tools for Neuro-Oncology: Connectomics and Spatial Transcriptomics - J. Germann, A. Boutet et al

Thu, 21 Apr 2022 20:00:17 GMT

Brain imaging, specifically magnetic resonance imaging (MRI), plays a key role in the clinical and research aspects of neuro-oncology. Novel neuroimaging techniques enable the transformation of a brain MRI into a so-called average brain. This allows projects using already acquired brain MRIs to perform group analyses and draw conclusions. Once the data are in this average brain, several types of analyses can be performed. For example, determining the most vulnerable locations for certain tumor types or perhaps even the underlying circuitry and gene expression that might cause predisposition to tumor growth. This information may further our understanding of tumor behavior, leading to better patient counseling, surgery timing, and treatment monitoring. - doi.org/10.3390/ cancers14030464 - 2022

PubReading [107] - Genome editing for Duchenne muscular dystrophy: a glimpse of the future? - C. Kupatt, M. Walter et al

Wed, 20 Apr 2022 20:00:17 GMT

Mutations in Dystrophin, one of the largest proteins in the mammalian body, are causative for a severe form of muscle disease, Duchenne Muscular Dystrophy (DMD), affecting not only skeletal muscle, but also the heart. In particular, exons 45–52 constitute a hotspot for DMD mutations. A variety of molecular therapies have been developed, comprising vectors encoding micro-and minidystrophins as well as utrophin, a protein with partially overlapping functions. With the advent of the CRISPR-Cas9-nuclease, genome editing offers a novel option of correction of the disease-causing mutations. Full restoration of the healthy gene by homology directed repair is a rare event. However, non-homologous end-joining (NHEJ) may restore the reading frame by causing exon excision. This approach has first been demonstrated in mice and then translated to large animals (dogs, pigs). This review discusses the potential opportunities and limitations of genome editing in DMD, including the generation of appropriate animal models as well as new developments in genome editing tools. - doi.org/10.1038/s41434-021-00222-4 - 2021

PubReading [106] - The Ultimate (Mis)match: When DNA Meets RNA - B. Palancade and R. Rothstein

Tue, 19 Apr 2022 20:00:16 GMT

RNA-containing structures, including ribonucleotide insertions, DNA:RNA hybrids and R-loops, have recently emerged as critical players in the maintenance of genome integrity. Strikingly, different enzymatic activities classically involved in genome maintenance contribute to their generation, their processing into genotoxic or repair intermediates, or their removal. Here we review how this substrate promiscuity can account for the detrimental and beneficial impacts of RNA insertions during genome metabolism. We summarize how in vivo and in vitro experiments support the contribution of DNA polymerases and homologous recombination proteins in the formation of RNA-containing structures, and we discuss the role of DNA repair enzymes in their removal. The diversity of pathways that are thus affected by RNA insertions likely reflects the ancestral function of RNA molecules in genome maintenance and transmission.

PubReading [104] - Structural principles of CRISPR-Cas enzymes used in nucleic acid detection - A. Dasa, H. Li et al

Fri, 15 Apr 2022 20:00:18 GMT

Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)-based technology has revolutionized the field of biomedicine with broad applications in genome editing, therapeutics and diagnostics. While a majority of applications involve the RNA-guided site-specific DNA or RNA cleavage by CRISPR enzymes, recent successes in nucleic acid detection rely on their collateral and non-specific cleavage activated by viral DNA or RNA. Ranging in enzyme composition, the mechanism for distinguishing self- from foreign-nucleic acids, the usage of second messengers, and enzymology, the CRISPR enzymes provide a diverse set of diagnosis tools in further innovations. Structural biology plays an important role in elucidating the mechanisms of these CRISPR enzymes. Here we summarize and compare structures of three types of CRISPR enzymes used in nucleic acid detection captured in their respective functional forms and illustrate the current understanding of their activation mechanism. - doi:10.1016/j.jsb.2022.107838 - 2022

PubReading [103] - IceBreaker: Software for high-resolution single-particle cryo-EM with non-uniform ice - M. Olek, P. Zhang et al

Thu, 14 Apr 2022 20:00:18 GMT

Despite the abundance of available software tools, optimal particle selection is still a vital issue in single-particle cryoelectron microscopy (cryo-EM). Regardless of the method used, most pickers struggle when ice thickness varies on a micrograph. IceBreaker allows users to estimate the relative ice gradient and flatten it by equalizing the local contrast. It allows the differentiation of particles from the background and improves overall particle picking performance. Furthermore, we introduce an additional parameter corresponding to local ice thickness for each particle. Particles with a defined ice thickness can be grouped and filtered based on this parameter during processing. These functionalities are especially valuable for on-the-fly processing to automatically pick as many particles as possible from each micrograph and to select optimal regions for data collection. Finally, estimated ice gradient distributions can be stored separately and used to inspect the quality of prepared samples. - doi.org/10.1016/j.str.2022.01.005 - 2022

PubReading [102] - Brain charts for the human lifespan - R. Bethlehem, J. Seidlitz, A. Alexander-Bloch et al

Wed, 13 Apr 2022 20:00:17 GMT

Over the past few decades, neuroimaging has become a ubiquitous tool in basic research and clinical studies of the human brain. However, no reference standards currently exist to quantify individual differences in neuroimaging metrics over time, in contrast to growth charts for anthropometric traits such as height and weight1. Here we assemble an interactive open resource to benchmark brain morphology derived from any current or future sample of MRI data (http://www.brainchart.io/). With the goal of basing these reference charts on the largest and most inclusive dataset available, acknowledging limitations due to known biases of MRI studies relative to the diversity of the global population, we aggregated 123,984 MRI scans, across more than 100 primary studies, from 101,457 human participants between 115 days post-conception to 100 years of age. MRI metrics were quantified by centile scores, relative to non-linear trajectories2 of brain structural changes, and rates of change, over the lifespan. Brain charts identified previously unreported neurodevelopmental milestones, showed high stability of individuals across longitudinal assessments, and demonstrated robustness to technical and methodological differences between primary studies. Centile scores showed increased heritability compared with non-centiled MRI phenotypes, and provided a standardized measure of atypical brain structure that revealed patterns of neuroanatomical variation across neurological and psychiatric disorders. In summary, brain charts are an essential step towards robust quantification of individual variation benchmarked to normative trajectories in multiple, commonly used neuroimaging phenotypes. - doi.org/10.1038/s41586-022-04554-y - 2022

PubReading [101] - Glycemic control in the critically ill: Less is more - G. Alhatemi, B. Seyoum et al

Tue, 12 Apr 2022 20:00:17 GMT

Hyperglycemia is associated with poor clinical outcomes in critically ill patients. Initial clinical trials of intensive insulin therapy targeting blood glucose levels of 80 to 110 mg/dL showed improved outcomes, but subsequent trials found no benefits and even increased harm with this approach. Emerging literature has evaluated other glycemic indices including time-in-target blood glucose range, glycemic variability, and stress hyperglycemia ratio. These indices, while well described in observational studies, have not been addressed in the initial trials. Additionally, the patient’s preexisting diabetes status and preadmission diabetic control may modulate the outcomes of stringent glycemic control, with worse outcomes of hyperglycemia being observed in patients without diabetes and in those with well-controlled diabetes. Most medical societies recommend less stringent glucose control in the range of 140 to 180 mg/dL for critically ill patients. - doi:10.3949/ccjm.89a.20171 - 2022

PubReading [100] - The complete sequence of a human genome - S. Nurk, S. Koren, A. Rhie, A. Phillips et al

Mon, 11 Apr 2022 20:00:18 GMT

Since its initial release in 2000, the human reference genome has covered only the euchromatic fraction of the genome, leaving important heterochromatic regions unfinished. Addressing the remaining 8% of the genome, the Telomere-to-Telomere (T2T) Consortium presents a complete 3.055 billion–base pair sequence of a human genome, T2T-CHM13, that includes gapless assemblies for all chromosomes except Y, corrects errors in the prior references, and introduces nearly 200 million base pairs of sequence containing 1956 gene predictions, 99 of which are predicted to be protein coding. The completed regions include all centromeric satellite arrays, recent segmental duplications, and the short arms of all five acrocentric chromosomes, unlocking these complex regions of the genome to variational and functional studies.- DOI: 10.1126/science.abj6987 - 2022

PubReading [99] - MicroED: conception, practice and future opportunities - M. Clabbers, A. Shiriaevaa and T. Gonen

Sun, 03 Apr 2022 20:00:18 GMT

This article documents a keynote seminar presented at the IUCr Congress in Prague, 2021. The cryo-EM method microcrystal electron diffraction is described and put in the context of macromolecular electron crystallography from its origins in 2D crystals of membrane proteins to today’s application to 3D crystals a millionth the size of that needed for X-ray crystallography. Milestones in method development and applications are described with an outlook to the future. - doi.org/10.1107/S2052252521013063 - 2022

PubReading [89] - Immunotherapy and prevention of pancreatic cancer - A. Morrison, K. Byrne and R. Vonderheide

Fri, 01 Apr 2022 20:00:17 GMT

Pancreatic cancer is the third leading cause of cancer mortality in the United States, recently surpassing breast cancer. A key component of pancreatic cancer’s lethality is its acquired immune privilege, which is driven by an immunosuppressive microenvironment, poor T-cell infiltration, and a low mutational burden. Although immunotherapies such as checkpoint blockade or engineered T cells have yet to demonstrate efficacy, a growing body of evidence suggests that orthogonal combinations of these and other strategies could unlock immunotherapy in pancreatic cancer. In this review, we will discuss promising immunotherapies currently under investigation in pancreatic cancer and provide a roadmap for the development of prevention vaccines for this and other cancers. - doi:10.1016/j.trecan.2018.04.001 - 2018

PubReading [97] - Ultra-High Dose Rate (FLASH) Radiotherapy: Silver Bullet or Fool’s Gold? - J. Wilson, K. Petersson et al.

Thu, 31 Mar 2022 20:00:15 GMT

Radiotherapy is a cornerstone of both curative and palliative cancer care. However, radiotherapy is severely limited by radiation-induced toxicities. If these toxicities could be reduced, a greater dose of radiation could be given therefore facilitating a better tumor response. Initial pre-clinical studies have shown that irradiation at dose rates far exceeding those currently used in clinical contexts reduce radiation-induced toxicities whilst maintaining an equivalent tumor response. This is known as the FLASH effect. To date, a single patient has been subjected to FLASH radiotherapy for the treatment of subcutaneous T-cell lymphoma resulting in complete response and minimal toxicities. The mechanism responsible for reduced tissue toxicity following FLASH radiotherapy is yet to be elucidated, but the most prominent hypothesis so far proposed is that acute oxygen depletion occurs within the irradiated tissue. This review examines the tissue response to FLASH radiotherapy, critically evaluates the evidence supporting hypotheses surrounding the biological basis of the FLASH effect, and considers the potential for FLASH radiotherapy to be translated into clinical contexts. - doi: 10.3389/fonc.2019.01563 - 2020

PubReading [96] - Highly accurate protein structure prediction for the human proteome - K. Tunyasuvunakool, D. Hassabis et al.

Wed, 30 Mar 2022 20:00:16 GMT

Protein structures can provide invaluable information, both for reasoning about biological processes and for enabling interventions such as structure-based drug development or targeted mutagenesis. After decades of effort, 17% of the total residues in human protein sequences are covered by an experimentally determined structure1. Here we markedly expand the structural coverage of the proteome by applying the state-of-the-art machine learning method, AlphaFold2, at a scale that covers almost the entire human proteome (98.5% of human proteins). The resulting dataset covers 58% of residues with a confident prediction, of which a subset (36% of all residues) have very high confidence. We introduce several metrics developed by building on the AlphaFold model and use them to interpret the dataset, identifying strong multi-domain predictions as well as regions that are likely to be disordered. Finally, we provide some case studies to illustrate how high-quality predictions could be used to generate biological hypotheses. We are making our predictions freely available to the community and anticipate that routine large-scale and high-accuracy structure prediction will become an important tool that will allow new questions to be addressed from a structural perspective.

PubReading [95] - A safe lithium mimetic for bipolar disorder - N. Singh, G. Churchill et al.

Tue, 29 Mar 2022 20:00:18 GMT

Lithium is the most effective mood stabilizer for the treatment of bipolar disorder, but it is toxic at only twice the therapeutic dosage and has many undesirable side effects. It is likely that a small molecule could be found with lithium-like efficacy but without toxicity through target-based drug discovery; however, therapeutic target of lithium remains equivocal. Inositol monophosphatase is a possible target but no bioavailable inhibitors exist. Here we report that the antioxidant ebselen inhibits inositol monophosphatase and induces lithium-like effects on mouse behaviour, which are reversed with inositol, consistent with a mechanism involving inhibition of inositol recycling. Ebselen is part of the National Institutes of Health Clinical Collection, a chemical library of bioavailable drugs considered clinically safe but without proven use. Therefore, ebselen represents a lithium mimetic with the potential both to validate inositol monophosphatase inhibition as a treatment for bipolar disorder and to serve as a treatment itself. - DOI: 10.1038/ncomms2320 - 2013

PubReading [94] - Tissue architecture in tumor initiation and progression - J. Almagro, A. Behrens

Mon, 28 Mar 2022 20:00:21 GMT

The 3D architecture of tissues bearing tumors impacts on the mechanical microenvironment of cancer, the accessibility of stromal cells, and the routes of invasion. A myriad of intrinsic and extrinsic forces exerted by the cancer cells, the host tissue, and the molecular and cellular microenvironment modulate the morphology of the tumor and its malignant potential through mechanical, biochemical, genetic, and epigenetic cues. Recent studies have investigated how tissue architecture influences cancer biology from tumor initiation and progression to distant metastatic seeding and response to therapy. With a focus on carcinoma, the most common type of cancer, this review discusses the latest discoveries on how tumor architecture is built and how tissue morphology affects the biology and progression of cancer cells.-doi:10.1016/j.trecan.2018.04.001 - 2018

PubReading [93] - Neuroprotective Actions of Dietary Choline - J. Blusztajn, B. Slack and T. Mellott

Fri, 25 Mar 2022 21:00:18 GMT

Choline is an essential nutrient for humans. It is a precursor of membrane phospholipids (e.g., phosphatidylcholine (PC)), the neurotransmitter acetylcholine, and via betaine, the methyl group donor S-adenosylmethionine. High choline intake during gestation and early postnatal development in rat and mouse models improves cognitive function in adulthood, prevents age-related memory decline, and protects the brain from the neuropathological changes associated with Alzheimer’s disease (AD), and neurological damage associated with epilepsy, fetal alcohol syndrome, and inherited conditions such as Down and Rett syndromes. These effects of choline are correlated with modifications in histone and DNA methylation in brain, and with alterations in the expression of genes that encode proteins important for learning and memory processing, suggesting a possible epigenomic mechanism of action. Dietary choline intake in the adult may also influence cognitive function via an effect on PC containing eicosapentaenoic and docosahexaenoic acids; polyunsaturated species of PC whose levels are reduced in brains from AD patients, and is associated with higher memory performance, and resistance to cognitive decline. - doi:10.3390/nu9080815 - 2017

PubReading [92] - Progress toward understanding chromosome silencing by Xist RNA - N. Brockdorff, J. Bowness, and G. Wei

Thu, 24 Mar 2022 21:36:26 GMT

The X inactive-specific transcript (Xist) gene is the master regulator of X chromosome inactivation in mammals. Xist produces a long noncoding (lnc)RNA that accumulates over the entire length of the chromosome from which it is transcribed, recruiting factors to modify under-lying chromatin and silence X-linked genes in cis. Recent years have seen significant progress in identifying important functional elements in Xist RNA, their associated RNA-binding proteins (RBPs), and the downstream pathways for chromatin modification and gene silencing. In this review, we summarize progress in understanding both how these pathways function in Xist-mediated silencing and the complex interplay between them. - doi/10.1101/gad.337196 - 2022

PubReading [91] - Circular Dichroism Spectroscopy of DNA: From Duplexes to Quadruplexes - M. Vorlickova, J. Kypr et al

Wed, 23 Mar 2022 20:58:12 GMT

Nucleic acids bear the genetic information and participate in its expression and evolution during replication, repair, recombination, transcription, and translation. These phenomena are mostly based on recognition of nucleic acids by proteins. The major factor enabling the specific recognition is structure. Circular dichroism (CD) spectroscopy is very useful to study secondary structures of nucleic acids, in general, and DNA, in particular. CD sensitively reflects isomerizations among distinct conformational states. The isomerizations may operate as molecular switches regulating various physiological or pathological processes. Here, we review CD spectra of nucleic acids, beginning with early studies on natural DNA molecules through analyses of synthetic polynucleotides to study of selected genomic fragments. - DOI: 10.1002/chir.22064 - 2012

PubReading [90] - Functional enzyme–polymer complexes - C. Waltman, M. Olvera de la Cruz

Tue, 22 Mar 2022 21:00:18 GMT

Engineered and native enzymes are poised to solve challenges in medicine, bioremediation, and biotechnology. One important goal is the possibility of upcycling polymers using enzymes. However, enzymes are often inactive in industrial, nonbiological conditions. It is particularly difficult to protect water-soluble enzymes at elevated temperatures by methods that preserve their functionality. Through atomistic and coarse-grained molecular dynamics simulations that capture protein conformational change, we show that an enzyme, PETase (polyethylene terephthalate [PET]), can be stabilized at elevated temperatures by complexation with random copolymers into nanoscale aggregates that do not precipitate into macroscopic phases. We demonstrated the efficiency of the method by simulating complexes of random copolymers and the enzyme PETase, which depolymerizes PET, a highly used polymer. These polymers are more industrially viable than peptides and can target specific domains on an enzyme. We design the mean composition of the random copolymers to control the polymer–enzyme surface contacts and the polymer conformation. When positioned on or near the active site, these polymer contacts can further stabilize the conformation of the active site at elevated temperatures. We explore the experimental implications of this active site stabilization method and show that PETase-random copolymer complexes have enhanced activity on both small molecule substrates and solid PET films. These results provide guidelines for engineering enzyme–polymer complexes with enhanced enzyme functionality in nonbiological environments. - doi.org/10.1073 - 2022

PubReading [89] - Novel Therapeutic Target(s) for Psoriatic Disease - V. Thakur and R. Mahajan

Mon, 21 Mar 2022 21:00:20 GMT

Psoriasis and psoriatic arthritis, together known as psoriatic disease, is highly prevalent chronic relapsing inflammatory disease affecting skin, joints or both and is associated with several comorbidities such as cardiovascular, metabolic, psychiatric, renal disease etc. The etiopathogenesis of psoriasis is complex and mainly driven by aberrant immune response owing to the genetic susceptibility and various environmental factors such as trauma, infections and drugs. Recent advances in understanding molecular and cellular pathways have identified tumor necrosis factor-α (TNF-α), interleukin-17 (IL-17), IL-23, IL-22 as major contributors in psoriasis pathogenesis. Advances in the knowledge of pathophysiology, the interaction of autoinflammation and clinical phenotypes have led to the development of highly effective targeted therapeutic agents which include TNF-α, IL-17, IL-23, IL-1 α/β or IL-36 inhibitors or receptor blockers, small molecule drugs like phosphodiesterase-4 inhibitors (apremilast), Janus kinase (JAK) inhibitors, retinoic acid receptor-related orphan receptor γt (RORγt) inhibitors. These novel drugs have promised the potential of improved disease control. In recent years, the transition from biologics to biosimilars especially with TNF-α inhibitors had significant impact on decreasing health care cost and increasing therapeutic options to the patients. However, selection of right treatment for an individual patient still remains challenging. Moreover, interplay between different epigenetic mechanisms such as the DNA methylation, chromatin modifications and noncoding RNA regulation has recently been started to be deciphered. Enzymes inhibitors involved in epigenetic pathways such as DNA methyltransferases and histone deacetylases demonstrated to restore normal epigenetic patterns in clinical settings and have provided the potential as novel therapeutic targets for psoriasis. In this review, we will discuss novel biologic agents and newer therapeutic approaches in treatment of psoriatic disease.

PubReading [88] - Factors influencing self-harm thoughts and behaviours over the first year of the COVID-19 pandemic in the UK: longitudinal analysis of 49 324 adults - E. Paul and D. Fancourt

Wed, 16 Mar 2022 23:27:35 GMT

There is concern that the COVID-19 pandemic and its aftermath will result in excess suicides by increasing known risk factors such as self-harm, but evidence on how pandemic-related risk factors contribute to changes in these outcomes is lacking.

In total, 26.1% and 7.9% of respondents reported self-harm thoughts and behaviours respectively at least once over the study period. The number of adverse experiences was more strongly related to outcomes than the number of worries. The largest specific adversity contributing to increases in both outcomes was having experienced physical or psychological abuse. Financial worries increased the likelihood of both outcomes in most age groups, and having had COVID-19 increased the likelihood of both outcomes in young (18–29 years) and middle-aged (45–59 years) adults. - 10.1192/bjp.2021.130 - 2022

PubReading [87] - Adipocytes, Innate Immunity and Obesity: A Mini-Review - A.Blaszczak, A. Jalilvand and W. Hsueh

Tue, 15 Mar 2022 21:00:19 GMT

The role of adipose tissue (AT) inflammation in obesity and its multiple related- complications is a rapidly expanding area of scientific interest. Within the last 30 years, the role of the adipocyte as an endocrine and immunologic cell has been progressively established. Like the macrophage, the adipocyte is capable of linking the innate and adaptive immune system through the secretion of adipokines and cytokines; exosome release of lipids, hormones, and microRNAs; and contact interaction with other immune cells. Key innate immune cells in AT include adipocytes, macrophages, neutrophils, and innate lymphoid cells type 2 (ILC2s). The role of the innate immune system in promoting adipose tissue inflammation in obesity will be highlighted in this review. T cells and B cells also play important roles in contributing to AT inflammation and are discussed in this series in the chapter on adaptive immunity. - doi: 10.3389/fimmu.2021.650768 - 2021

PubReading [86] - What is the structural chemistry of the living organism at its temperature and pressure? - J. Helliwell

Mon, 14 Mar 2022 21:00:19 GMT

The three probes of the structure of matter (X-rays, neutrons and electrons) in biology have complementary properties and strengths. The balance between these three probes within their strengths and weaknesses is perceived to change, even dramatically so at times. For the study of combined states of order and disorder, NMR crystallography is also applicable. Of course, to understand biological systems the required perspectives are surely physiologically relevant temperatures and relevant chemical conditions, as well as a minimal perturbation owing to the needs of the probe itself. These remain very tough challenges because, for example, cryoEM by its very nature will never be performed at room temperature, crystallization often requires nonphysiological chemical conditions, and X-rays and electrons cause beam damage. However, integrated structural biology techniques and functional assays provide a package towards physiological relevance of any given study. Reporting of protein crystal structures, and their associated database entries, could usefully indicate how close to the biological situation they are, as discussed in detail in this feature article. - doi.org/10.1107/S2059798320000546 - 2020

PubReading [85] - Low-dose metformin targets the lysosomal AMPK pathway through PEN2 - T. Ma, S. Lin

Mon, 07 Mar 2022 21:33:44 GMT

Metformin, the most prescribed antidiabetic medicine, has shown other benefits such as anti-ageing and anticancer effects1–4. For clinical doses of metformin, AMP-activated protein kinase (AMPK) has a major role in its mechanism of action4,5; however, the direct molecular target of metformin remains unknown. Here we show that clinically relevant concentrations of metformin inhibit the lysosomal proton pump v-ATPase, which is a central node for AMPK activation following glucose starvation6. We synthesize a photoactive metformin probe and identify PEN2, a subunit of γ-secretase7, as a binding partner of metformin with a dissociation constant at micromolar levels. Metformin-bound PEN2 forms a complex with ATP6AP1, a subunit of the v-ATPase8, which leads to the inhibition of v-ATPase and the activation of AMPK without effects on cellular AMP levels. Knockout of PEN2 or re-introduction of a PEN2 mutant that does not bind ATP6AP1 blunts AMPK activation. In vivo, liver-specific knockout of Pen2 abolishes metformin-mediated reduction of hepatic fat content, whereas intestine-specific knockout of Pen2 impairs its glucose-lowering effects. Furthermore, knockdown of pen-2 in Caenorhabditis elegans abrogates metformin- induced extension of lifespan. Together, these findings reveal that metformin binds PEN2 and initiates a signalling route that intersects, through ATP6AP1, the lysosomal glucose-sensing pathway for AMPK activation. This ensures that metformin exerts its therapeutic benefits in patients without substantial adverse effects. - https://doi.org/10.1038/s41586-022-04431-8 - 2022

PubReading [84] - Neurotrauma and Repair Research: Traumatic Brain Injury (TBI) and its Treatments - H. Algattas and J. Huang

Tue, 01 Mar 2022 21:00:19 GMT

Traumatic brain injury (TBI) affects a growing portion of the population and continues to take national spotlight with advances in imaging technology and understanding of long-term effects. However, there is large variance in TBI treatment protocols due to injury variability and lack of both mechanistic understanding and strong treatment recommendations. Recent practice suggests three disparate treatment approaches, all which aim at promoting neuroprotection after TBI, show promise: immediate hypothermia, hyper- baric oxygen, and progesterone supplementation. The research is controversial at times, yet there are abundant opportunities to develop the technology behind hypothermia and hyperbaric oxygen treatments which would surely aid in aligning the current data. Additionally, while progesterone has already been packaged in nanoparticle form it may benefit from continued formulation and administration research. The treatments and the avenues for improvement are reviewed in the present paper. - 10.4137/BECB.S10968 - 2013

PubReading [83] - CAR T cells produced in vivo to treat cardiac injury - J. Rurik, J. Epstein et al.

Thu, 13 Jan 2022 21:00:18 GMT

Fibrosis affects millions of people with cardiac disease. We developed a therapeutic approach to generate transient antifibrotic chimeric antigen receptor (CAR) T cells in vivo by delivering modified messenger RNA (mRNA) in T cell–targeted lipid nanoparticles (LNPs). The efficacy of these in vivo–reprogrammed CAR T cells was evaluated by injecting CD5-targeted LNPs into a mouse model of heart failure. Efficient delivery of modified mRNA encoding the CAR to T lymphocytes was observed, which produced transient, effective CAR T cells in vivo. Antifibrotic CAR T cells exhibited trogocytosis and retained the target antigen as they accumulated in the spleen. Treatment with modified mRNA-targeted LNPs reduced fibrosis and restored cardiac function after injury. In vivo generation of CAR T cells may hold promise as a therapeutic platform to treat various diseases. - DOI: 10.1126/science.abm0594 - 2022

PubReading [82] - How to Write a Systematic Review - R. Wright, K. Spindler et al.

Wed, 12 Jan 2022 21:00:19 GMT

Evidence-based medicine (EBM) is the combination of the best available research evidence with clinical experience and patient needs. The concept of EBM as a part of clinical decision making has become increasingly popular over the last decade. In the hierarchy of studies meta-analysis and systematic reviews occupy the highest levels. A systematic review of a clinical question can be performed by following a relatively standard form. These techniques as described here can be performed without formal training. Systematic reviews conducted in this fashion can be used as a higher form of current concepts or as review articles and replace the traditional expert opinion narrative review. - DOI: 10.1097/BLO.0b013e31802c9098

PubReading [81] - The N-terminal cysteine is a dual sensor of oxygen and oxidative stress - A. Jung Heo, Y. Tae Kwon et al.

Mon, 20 Dec 2021 22:02:33 GMT

Cellular homeostasis requires the sensing of and adaptation to intracellular oxygen (O2) and reactive oxygen species (ROS). The Arg/N-degron pathway targets proteins that bear destabilizing N-terminal residues for degradation by the proteasome or via autophagy. Under normoxic conditions, the N-terminal Cys (Nt-Cys) residues of specific substrates can be oxidized by dioxygenases such as plant cysteine oxidases and cysteamine (2-aminoethane- thiol) dioxygenases and arginylated by ATE1 R-transferases to generate Arg-CysO2(H) (R-CO2). Proteins bearing the R-CO2 N-degron are targeted via Lys48 (K48)–linked ubiquitylation by UBR1/UBR2 N-recognins for proteasomal degradation. During acute hypoxia, such proteins are partially stabilized, owing to decreased Nt-Cys oxidation. Here, we show that if hypoxia is prolonged, the Nt-Cys of regulatory proteins can be chemically oxidized by ROS to generate Arg-CysO3(H) (R-CO3), a lysosomal N-degron. The resulting R-CO3 is bound by KCMF1, a N-recognin that induces K63-linked ubiquitylation, followed by K27-linked ubiquitylation by the noncanonical N-recognin UBR4. Autophagic targeting of Cys/N-degron substrates is mediated by the autophagic N-recognin p62/SQTSM- 1/Sequestosome-1 through recognition of K27/K63-linked ubiquitin (Ub) chains. This Cys/N-degron–dependent reprogramming in the proteolytic flux is important for cellular homeostasis under both chronic hypoxia and oxidative stress. A small-compound ligand of p62 is cytoprotective under oxidative stress through its ability to accelerate proteolytic flux of K27/K63-ubiquitylated Cys/N-degron substrates. Our results suggest that the Nt-Cys of conditional Cys/ N-degron substrates acts as an acceptor of O2 to maintain both O2 and ROS homeostasis and modulates half-lives of substrates through either the proteasome or lysosome by reprogramming of their Ub codes. - https://doi.org/10.1073/pnas.2107993118 - 2021

PubReading [80] - Exercise plasma boosts memory and dampens brain inflammation via clusterin - Z. De Miguel, T. Wyss-Coray et al.

Tue, 21 Dec 2021 21:00:20 GMT

Physical exercise is generally beneficial to all aspects of human and animal health, slowing cognitive ageing and neurodegeneration1. The cognitive benefits of physical exercise are tied to an increased plasticity and reduced inflammation within the hippocampus2–4, yet little is known about the factors and mechanisms that mediate these effects. Here we show that ‘runner plasma’, collected from voluntarily running mice and infused into sedentary mice, reduces baseline neuroinflammatory gene expression and experimentally induced brain inflammation. Plasma proteomic analysis revealed a concerted increase in complement cascade inhibitors including clusterin (CLU). Intravenously injected CLU binds to brain endothelial cells and reduces neuroinflammatory gene expression in a mouse model of acute brain inflammation and a mouse model of Alzheimer’s disease. Patients with cognitive impairment who participated in structured exercise for 6 months had higher plasma levels of CLU. These findings demonstrate the existence of anti-inflammatory exercise factors that are transferrable, target the cerebrovasculature and benefit the brain, and are present in humans who engage in exercise. - https://doi.org/10.1038/s41586-021-04183-x - 2021

PubReading [79] - Nanopore sequencing technology, bioinformatics and applications - Y. Wang, K. Fai Au et al.

Mon, 20 Dec 2021 21:40:54 GMT

Rapid advances in nanopore technologies for sequencing single long DNA and RNA molecules have led to substantial improvements in accuracy, read length and throughput. These breakthroughs have required extensive development of experimental and bioinformatics methods to fully exploit nanopore long reads for investigations of genomes, transcriptomes, epigenomes and epitranscriptomes. Nanopore sequencing is being applied in genome assembly, full-length transcript detection and base modification detection and in more specialized areas, such as rapid clinical diagnoses and outbreak surveillance. Many opportunities remain for improving data quality and analytical approaches through the development of new nanopores, base-calling methods and experimental protocols tailored to particular applications. - https://doi.org/10.1038/s41587-021-01108-x - 2021

PubReading [78] - Megahertz serial crystallography - Max Wiedorn et al.

Tue, 14 Dec 2021 21:00:18 GMT

The new European X-ray Free-Electron Laser is the first X-ray free-electron laser capable of delivering X-ray pulses with a megahertz inter-pulse spacing, more than four orders of magnitude higher than previously possible. However, to date, it has been unclear whether it would indeed be possible to measure high-quality diffraction data at megahertz pulse repetition rates. Here, we show that high-quality structures can indeed be obtained using currently available operating conditions at the European XFEL. We present two complete data sets, one from the well-known model system lysozyme and the other from a so far unknown complex of a β-lactamase from K. pneumoniae involved in antibiotic resistance. This result opens up megahertz serial femtosecond crystallography (SFX) as a tool for reliable structure determination, substrate screening and the efficient measurement of the evolution and dynamics of molecular structures using megahertz repetition rate pulses available at this new class of X-ray laser source. - DOI: 10.1038/s41467-018-06156-7 - 2018

PubReading [77] - Engineering transplantable jejunal mucosal grafts using patient- derived organoids from children with intestinal failure - M. Laween, V. Li et al.

Mon, 13 Dec 2021 21:33:26 GMT

Intestinal failure (IF), following extensive anatomical or functional loss of small intestine (SI), has debilitating long-term consequences on children1. The priority of patient care is to increase the length of functional intestine, particularly the jejunum, to promote nutritional independence2. Here we construct autologous jejunal mucosal grafts using pediatric patient biomaterials and show that patient-derived organoids (PDO) can be expanded efficiently in vitro. In parallel, we generate decellularized human intestinal matrix with intact nanotopography, that forms biological scaffolds. Proteomic and Raman spectroscopy analyses reveal highly analogous biochemical profiles of human SI and colon scaffolds, indicating that they can be used interchangeably as platforms for intestinal engineering. Indeed, seeding of jejunal organoids onto either type of scaffold reliably reconstructs grafts that exhibit several aspects of physiological jejunal function, and that survive to form luminal structures after transplantation into the kidney capsule or subcutaneous pockets for up to 2 weeks. Our findings provide proof-of-concept data for engineering patient-specific jejunal grafts for children with IF, ultimately aiding in the restoration of nutritional autonomy. - doi:10.1038/s41591-020-1024-z. - 2020

PubReading [76] - The Catalytic Asymmetric Intermolecular Prins Reaction - D. Díaz-Oviedo, R. Maji, and B. List

Fri, 10 Dec 2021 21:00:20 GMT

Despite their significant potential, catalytic asymmetric reactions of olefins with formaldehyde are rare and metal-free approaches have not been previously disclosed. Here we describe an enantioselective intermolecular Prins reaction of styrenes and paraformaldehyde to form 1,3-dioxanes, using confined imino-imidodiphosphate (iIDP) Brønsted acid catalysts. Isotope labeling experiments and computations suggest a concerted, highly asynchronous addition of an acid-activated formaldehyde oligomer to the olefin. The enantioenriched 1,3-dioxanes can be transformed into the corresponding optically active 1,3-diols, which are valuable synthetic building blocks. - https://doi.org/10.1021/jacs.1c10245 - 2021

PubReading [74] - Sodium/glucose cotransporter 2 (SGLT2) inhibitors improve cardiac function by reducing JunD expression in human diabetic hearts - R. Marfella, G. Paolisso et al.

Thu, 09 Dec 2021 21:00:22 GMT

Background: The pathogenesis of experimental diabetic cardiomyopathy may involve the activator protein 1 (AP-1) member, JunD. Using non-diabetic heart transplant (HTX) in recipients with diabetes, we examined the effects of the diabetic milieu (hyperglycemia and insulin resistance) on cardiac JunD expression over 12 months. Because sodium/glucose cotransporter-2 inhibitors (SGLT2i) significantly reverse high glucose-induced AP-1 binding in the proximal tubular cell, we investigated JunD expression in a subgroup of type 2 diabetic recipients receiving SGLT2i treatment.

Methods: We evaluated 77 first HTX recipients (40 and 37 patients with and without diabetes, respectively). Among the recipients with diabetes, 17 (45.9%) were receiving SGLT2i treatment. HTX recipients underwent standard clinical evaluation (metabolic status, echocardiography, coronary computed tomography angiography, and endomyocardial biopsy). In the biopsy samples, we evaluated JunD, insulin receptor substrates 1 and 2 (IRS1 and IRS2), peroxisome proliferator-activated receptor-γ (PPAR-γ), and ceramide levels using real-time polymerase chain reaction and immunofluorescence. The biopsy evaluations in this study were performed at 1–4 weeks (basal), 5–12 weeks (intermediate), and up to 48 weeks (final, end of 12-month follow-up) after HTX.

Results: There was a significant early and progressive increase in the cardiac expression of JunD/PPAR-γ and ceramide levels, along with a significant decrease in IRS1 and IRS2 in recipients with diabetes but not in those without diabetes. These molecular changes were blunted in patients with diabetes receiving SGLT2i treatment. Conclusion: Early pathogenesis in human diabetic cardiomyopathy is associated with JunD/PPAR-γ overexpression and lipid accumulation following HTX in recipients with diabetes. Remarkably, this phenomenon was reduced by concomitant therapy with SGLT2i, which acted directly on diabetic hearts. - doi.org/10.1016/j.metabol.2021.154936 - 2021

PubReading [73] - Electronic health records and polygenic risk scores for predicting disease risk - R. Li, J. Moore et al.

Wed, 08 Dec 2021 21:00:15 GMT

Accurate prediction of disease risk based on the genetic make-up of an individual is essential for effective prevention and personalized treatment. Nevertheless, to date, individual genetic variants from genome-wide association studies have achieved only moderate prediction of disease risk. The aggregation of genetic variants under a polygenic model shows promising improvements in prediction accuracies. Increasingly, electronic health records (EHRs) are being linked to patient genetic data in biobanks, which provides new opportunities for developing and applying polygenic risk scores in the clinic, to systematically examine and evaluate patient susceptibilities to disease. However, the heterogeneous nature of EHR data brings forth many practical challenges along every step of designing and implementing risk prediction strategies. In this Review, we present the unique considerations for using genotype and phenotype data from biobank-linked EHRs for polygenic risk prediction. - doi.org/10.1038/s41576-020-0224-1 - 2020

PubReading [72] - Delivery strategies for macromolecular drugs in cancer therapy - Q. Guo, C. Jiang et al.

Tue, 07 Dec 2021 21:00:15 GMT

With the development of biotherapy, biomacromolecular drugs have gained tremendous attention recently, especially in drug development field due to the sophisticated functions in vivo. Over the past few years, a motley variety of drug delivery strategies have been developed for bio- macromolecular drugs to overcome the difficulties in the druggability, e.g., the instability and easily restricted by physiologic barriers. The application of novel delivery systems to deliver bio- macromolecular drugs can usually prolong the half-life, increase the bioavailability, or improve patient compliance, which greatly improves the efficacy and potentiality for clinical use of biomacromolecular drugs. In this review, recent studies regarding the drug delivery strategies for macromolecular drugs in cancer therapy are summarized, mainly drawing on the development over the last five years. - doi.org/10.1016/j.apsb.2020.01.009 - 2019

PubReading [71] - Optical Microscopy and Electron Microscopy for the Morphological Evaluation of Tendons: A Mini Review - M. Xu, B. Liu et al.

Mon, 06 Dec 2021 21:00:15 GMT

The morphological characteristics of tendons have been thoroughly evaluated via microscopy. Optical microscopy and electron microscopy are the most commonly used techniques for tendon tissue observation. According to the principles of both microscopy types, preparation and evaluation methods vary. Simple optical microscopy is commonly used in the observation of cells and extracellular matrix, and many stains, including hematoxylineosin, Van Gieson, Prussian blue, Alcian blue, and toluidine blue, are used for evaluating cells, collagen fiber arrangement, and non-collagenous proteins. Histological scoring systems have been used in many studies for semi-quantification. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) are the most commonly used electron microscopy types, and special consideration is needed for the fixation and embedding protocols. Glutaraldehyde followed by osmium is most commonly used in the chemical fixation of tendon tissue, followed by epoxy resin embedment. Longitudinal sections captured in SEM images show the arrangement of collagen fibrils and the cells and lipid drops among them, while cross-sections captured in TEM images show the diameter and distribution of collagen fibrils. SEM and TEM are used together for comprehensive evaluations. This mini review is focused on the preparation methodology and related evaluation indexes for the morphological evaluation of tendons. - DOI: 10.1111/os.12637 - 2020

PubReading [70] - Flavonoids as Anticancer Agents - D. Kopustinskiene, J. Bernatoniene et al

Sat, 04 Dec 2021 21:00:15 GMT

Flavonoids are polyphenolic compounds subdivided into 6 groups: isoflavonoids, flavanones, flavanols, flavonols, flavones and anthocyanidins found in a variety of plants. Fruits, vegetables, plant-derived beverages such as green tea, wine and cocoa-based products are the main dietary sources of flavonoids. Flavonoids have been shown to possess a wide variety of anticancer effects: they modulate reactive oxygen species (ROS)-scavenging enzyme activities, participate in arresting the cell cycle, induce apoptosis, autophagy, and suppress cancer cell proliferation and invasiveness. Flavonoids have dual action regarding ROS homeostasis—they act as antioxidants under normal conditions and are potent pro-oxidants in cancer cells triggering the apoptotic pathways and downregulating pro-inflammatory signaling pathways. This article reviews the biochemical properties and bioavailability of flavonoids, their anticancer activity and its mechanisms of action. - doi:10.3390/nu12020457 - 2020

PubReading [69]- Targeting BMI-1 in B cells restores effective humoral immune responses and controls chronic viral infection - A. Di Pietro, K. Good-Jacobson et al

Fri, 03 Dec 2021 21:00:15 GMT

Ineffective antibody-mediated responses are a key characteristic of chronic viral infection. However, our understanding of the intrinsic mechanisms that drive this dysregulation are unclear. Here, we identify that targeting the epigenetic modifier BMI-1 in mice improves humoral responses to chronic lymphocytic choriomeningitis virus. BMI-1 was upregulated by germinal center B cells in chronic viral infection, correlating with changes to the accessible chromatin landscape, compared to acute infection. B cell-intrinsic deletion of Bmi1 accelerated viral clearance, reduced splenomegaly and restored splenic architecture. Deletion of Bmi1 restored c-Myc expression in B cells, concomitant with improved quality of antibody and coupled with reduced antibody-secreting cell numbers. Specifically, BMI-1-deficiency induced antibody with increased neutralizing capacity and enhanced antibody-dependent effector function. Using a small molecule inhibitor to murine BMI-1, we could deplete antibody-secreting cells and prohibit detrimental immune complex formation in vivo. This study defines BMI-1 as a crucial immune modifier that controls antibody-mediated responses in chronic infection. - doi.org/10.1038/s41590-021-01077-y - 2021

PubReading [68] - Omega-3 fatty acids attenuate dendritic cell function via NF-κB independent of PPARγ - E. Draper and H. Roche

Thu, 02 Dec 2021 21:00:16 GMT

Long-chain n-3 polyunsaturated fatty acids (n-3 PUFA) have been shown to modulate the immune response and have therapeutic effects in inflammatory disorders. PUFA are also peroxisome proliferators-activator receptor-gamma (PPARγ) ligands; a family of ligand-activated transcription factors, which when activated antagonise the pro-inflammatory capability of nuclear factor κB (NF-κB). PPARγ plays a role in dendritic cell (DC) maturation and n-3 PUFA have been shown to affect DC maturation by decreasing activation of NF-κB. While n-3 PUFA can function as PPAR ligands, it is not known whether the NF-κB-mediated immunomodulatory properties of n-3 PUFA are PPARγ-dependent. In this study we examined whether the immunomodulatory effects of n-3 PUFA on DC activation were mediated through activation of PPARγ. Treatment of murine bone marrow derived DCs with docosahexaenoic acid (DHA; 25 μM) and eicosapentaenoic acid (EPA; 25 μM) attenuated LPS-induced DC maturation. This was characterised by suppression of IL-12 production and expression of CD40, CD80, CD86 and MHC II and enhanced production of IL-10 and expression of IL-10R. This was coincident with enhanced PPARγ expression, suppressed NF-κB activity and increased the physical interaction and cellular colocalization between NF-κB with PPARγ. To understand the functional implication of the physical association of PPARγ with NF-κB, we determined whether the specific PPARγ inhibitor, GW9662 could abolish the anti-inflammatory effect of n-3 PUFA Inhibiting PPARγ did not impede the NF-κB-mediated anti-inflammatory cytokine profile induced by EPA and DHA alone. Thus n-3 PUFA activate PPARγ and interact with NF-κB in DC. However, the anti-inflammatory effects of EPA and DHA on DCs are independent of PPARγ. - 10.1016/j.jnutbio.2010.06.009 - 2011

PubReading [67] - DNA methylation in human sperm: a systematic review - F. Asenius, A. Danson and S. Marzi

Wed, 01 Dec 2021 21:00:15 GMT

BACKGROUND: Studies in non-human mammals suggest that environmental factors can influence spermatozoal DNA methylation, and some research suggests that spermatozoal DNA methylation is also implicated in conditions such as subfertility and imprinting disorders in the offspring. Together with an increased availability of cost-effective methods of interrogating DNA methylation, this premise has led to an increasing number of studies investigating the DNA methylation landscape of human spermatozoa. However, how the human spermatozoal DNA methylome is influenced by environmental factors is still unclear, as is the role of human spermatozoal DNA methylation in subfertility and in influencing offspring health.

OBJECTIVE AND RATIONALE: The aim of this systematic review was to critically appraise the quality of the current body of literature on DNA methylation in human spermatozoa, summarize current knowledge and generate recommendations for future research.

SEARCH METHODS: A comprehensive literature search of the PubMed, Web of Science and Cochrane Library databases was conducted using the search terms ‘semen’ OR ‘sperm’ AND ‘DNA methylation’. Publications from 1 January 2003 to 2 March 2020 that studied human sperm and were written in English were included. Studies that used sperm DNA methylation to develop methodologies or forensically identify semen were excluded, as were reviews, commentaries, meta-analyses or editorial texts. The Grading of Recommendations, Assessment, Development and Evaluations (GRADE) criteria were used to objectively evaluate quality of evidence in each included publication.

OUTCOMES: The search identified 446 records, of which 135 were included in the systematic review. These 135 studies were divided into three groups according to area of research; 56 studies investigated the influence of spermatozoal DNA methylation on male fertility and abnormal semen parameters, 20 studies investigated spermatozoal DNA methylation in pregnancy outcomes including offspring health and 59 studies assessed the influence of environmental factors on spermatozoal DNA methylation. Findings from studies that scored as ‘high’ and ‘moderate’ quality of evidence according to GRADE criteria were summarized. We found that male subfertility and abnormal semen parameters, in particular oligozoospermia, appear to be associated with abnormal spermatozoal DNA methylation of imprinted regions. However, no specific DNA methylation signature of either subfertility or abnormal semen parameters has been convincingly replicated in genome-scale, unbiased analyses. Furthermore, although findings require independent replication, current evidence suggests that the spermatozoal DNA methylome is influenced by cigarette smoking, advanced age and environmental pollutants. Importantly however, from a clinical point of view, there is no convincing evidence that changes in spermatozoal DNA methylation influence pregnancy outcomes or offspring health.

WIDER IMPLICATIONS: Although it appears that the human sperm DNA methylome can be influenced by certain environmental and physiological traits, no findings have been robustly replicated between studies. We have generated a set of recommendations that would enhance the reliability and robustness of findings of future analyses of the human sperm methylome. Such studies will likely require multicentre collaborations to reach appropriate sample sizes, and should incorporate phenotype data in more complex statistical models.

doi:10.1093/humupd/dmaa025 - 2020

PubReading [66] - The 2021 Nucleic Acids Research database issue and the online molecular biology database collection - D. Rigden and X. Fernandez

Tue, 30 Nov 2021 21:00:15 GMT

The 2021 Nucleic Acids Research database Issue contains 189 papers spanning a wide range of biological fields and investigation. It includes 89 papers reporting on new databases and 90 covering recent changes to resources previously published in the Issue. A further ten are updates on databases most recently published elsewhere. Seven new databases focus on COVID-19 and SARS-CoV- 2 and many others offer resources for studying the virus. Major returning nucleic acid databases include NONCODE, Rfam and RNAcentral. Protein family and domain databases include COG, Pfam, SMART and Panther. Protein structures are covered by RCSB PDB and dispersed proteins by PED and MobiDB. In metabolism and signalling, STRING, KEGG and WikiPathways are featured, along with returning KLIFS and new DKK and KinaseMD, all focused on kinases. IMG/M and IMG/VR update in the microbial and viral genome resources section, while human and model organism genomics resources include Flybase, Ensembl and UCSC Genome Browser. Cancer studies are covered by updates from canSAR and PINA, as well as newcomers CNCdatabase and Oncovar for cancer drivers. Plant comparative genomics is catered for by updates from Gramene and GreenPhylDB. The entire Database Issue is freely available online on the Nucleic Acids Research website (https://academic.oup.com/nar). The NAR online Molecular Biology Database Collection has been substantially updated, revisiting nearly 1000 entries, adding 90 new resources and eliminating 86 obsolete databases, bringing the current total to 1641 databases. It is available at https://www. oxfordjournals.org/nar/database/c/. - doi: 10.1093/nar/gkaa1216 - 2021

PubReading [65] - Cell Cycle Regulation of Stem Cells by MicroRNAs - M. Mens and M. Ghanbari

Mon, 29 Nov 2021 21:00:16 GMT

MicroRNAs (miRNAs) are a class of small non-coding RNA molecules involved in the regulation of gene expression. They are involved in the fine-tuning of fundamental biological processes such as proliferation, differentiation, survival and apoptosis in many cell types. Emerging evidence suggests that miRNAs regulate critical pathways involved in stem cell function. Several miRNAs have been suggested to target transcripts that directly or indirectly coordinate the cell cycle progression of stem cells. Moreover, previous studies have shown that altered expression levels of miRNAs can contribute to pathological conditions, such as cancer, due to the loss of cell cycle regulation. However, the precise mechanism underlying miRNA- mediated regulation of cell cycle in stem cells is still incompletely understood. In this review, we discuss current knowledge of miRNAs regulatory role in cell cycle progression of stem cells. We describe how specific miRNAs may control cell cycle associated molecules and checkpoints in embryonic, somatic and cancer stem cells. We further outline how these miRNAs could be regulated to influence cell cycle progression in stem cells as a potential clinical application. - doi.org/10.1007/s12015-018-9808-y - 2018

PubReading [64] - Lysosome-targeting chimaeras for degradation of extracellular proteins - S. Banik, C. Bertozzi et al

Wed, 24 Nov 2021 21:00:16 GMT

The majority of therapies that target individual proteins rely on specific activity- modulating interactions with the target protein—for example, enzyme inhibition or ligand blocking. However, several major classes of therapeutically relevant proteins have unknown or inaccessible activity profiles and so cannot be targeted by such strategies. Protein-degradation platforms such as proteolysis-targeting chimaeras (PROTACs)1,2 and others (for example, dTAGs3, Trim-Away4, chaperone-mediated autophagy targeting5 and SNIPERs6) have been developed for proteins that are typically difficult to target; however, these methods involve the manipulation of intracellular protein degradation machinery and are therefore fundamentally limited to proteins that contain cytosolic domains to which ligands can bind and recruit the requisite cellular components. Extracellular and membrane-associated proteins—the products of 40% of all protein-encoding genes7—are key agents in cancer, ageing-related diseases and autoimmune disorders8, and so a general strategy to selectively degrade these proteins has the potential to improve human health. Here we establish the targeted degradation of extracellular and membrane-associated proteins using conjugates that bind both a cell-surface lysosome-shuttling receptor and the extracellular domain of a target protein. These initial lysosome-targeting chimaeras, which we term LYTACs, consist of a small molecule or antibody fused to chemically synthesized glycopeptide ligands that are agonists of the cation-independent mannose-6-phosphate receptor (CI-M6PR). We use LYTACs to develop a CRISPR interference screen that reveals the biochemical pathway for CI-M6PR-mediated cargo internalization in cell lines, and uncover the exocyst complex as a previously unidentified—but essential—component of this pathway. We demonstrate the scope of this platform through the degradation of therapeutically relevant proteins, including apolipoprotein E4, epidermal growth factor receptor, CD71 and programmed death-ligand 1. Our results establish a modular strategy for directing secreted and membrane proteins for lysosomal degradation, with broad implications for biochemical research and for therapeutics. - doi.org/10.1038/s41586-020-2545-9 - 2020

PubReading [63] - Integrated analysis of multimodal single-cell data -Y. Hao, R. Satija

Tue, 23 Nov 2021 21:00:15 GMT

The simultaneous measurement of multiple modalities represents an exciting frontier for single-cell genomics and necessitates computational methods that can define cellular states based on multimodal data. Here, we introduce ‘‘weighted-nearest neighbor’’ analysis, an unsupervised framework to learn the relative utility of each data type in each cell, enabling an integrative analysis of multiple modalities. We apply our procedure to a CITE-seq dataset of 211,000 human peripheral blood mononuclear cells (PBMCs) with panels extending to 228 antibodies to construct a multimodal reference atlas of the circulating immune system. Multimodal analysis substantially improves our ability to resolve cell states, allowing us to identify and validate previously unreported lymphoid subpopulations. Moreover, we demonstrate how to leverage this reference to rapidly map new datasets and to interpret immune responses to vaccination and coronavirus disease 2019 (COVID-19). Our approach represents a broadly applicable strategy to analyze single-cell multimodal data- sets and to look beyond the transcriptome toward a unified and multimodal definition of cellular identity. - doi.org/10.1016/j.cell.2021.04.048 - 2021

PubReading [62] - Molecular Printing with DNA Nanotechnology - F. Rizzuto, T. Trinh and H. Sleiman

Mon, 22 Nov 2021 21:00:15 GMT

DNA nanotechnology has generated a wealth of structures uniquely suited for nanoscale patterning; however, scalability, affordability, and recyclability are important preconditions for the industrial production and widespread use of DNA-based materials. In this perspective, we propose that ‘‘printing’’ programmed particles from transient DNA templates provides a practical pathway toward overcoming these hurdles. Just as a printing press transfers ink to paper in a cyclical process, DNA can translocate materials from one substrate to another while preserving spatial information. DNA printing decorates a particle with arbitrarily designed patterns that break its symmetry, enabling its autonomous assembly into complex structures. Printing thus organizes anisotropic particles with high throughput: a single DNA scaffold engenders template recyclability and hybrid material scale-up. By applying DNA-based printing methods to increasingly diverse materials, we will export DNA nanotechnology to other research areas and generate multivalent, asymmetric structures for scalable, nanoscale control of structure and function. - doi.org/10.1016/j.chempr.2020.06.012 - 2020

PubReading [61] - Discovery of a novel pseudo β-hairpin structure of N-truncated amyloid-β for use as a vaccine against Alzheimer’s disease - P. Bakrania, T. Bayer et al

Thu, 18 Nov 2021 21:00:16 GMT

One of the hallmarks of Alzheimer’s disease (AD) are deposits of amyloid-beta (Aβ) protein in amyloid plaques in the brain. The Aβ peptide exists in several forms, including full-length Aβ1-42 and Aβ1-40 – and the N-truncated species, pyroglutamate Aβ3-42 and Aβ4-42, which appear to play a major role in neurodegeneration. We previously identified a murine antibody (TAP01), which binds specifically to soluble, non-plaque N-truncated Aβ species. By solving crystal structures for TAP01 family antibodies bound to pyroglutamate Aβ3-14, we identified a novel pseudo β-hairpin structure in the N-terminal region of Aβ and show that this underpins its unique binding properties. We engineered a stabilised cyclic form of Aβ1-14 (N-Truncated Amyloid Peptide AntibodieS; the ‘TAPAS’ vaccine) and showed that this adopts the same 3-dimensional conformation as the native sequence when bound to TAP01. Active immunisation of two mouse models of AD with the TAPAS vaccine led to a striking reduction in amyloid- plaque formation, a rescue of brain glucose metabolism, a stabilisation in neuron loss, and a rescue of memory deficiencies. Treating both models with the humanised version of the TAP01 antibody had similar positive effects. Here we report the discovery of a unique conformational epitope in the N-terminal region of Aβ, which offers new routes for active and passive immunisation against AD. - doi.org/10.1038/s41380-021-01385-7. -2021

PubReading [60] - DNA helicases in homologous recombination repair - D. Branzei and B. Szakal

Wed, 17 Nov 2021 21:00:17 GMT

Helicases are in the spotlight of DNA metabolism and are critical for DNA repair in all domains of life. At their biochemical core, they bind and hydrolyze ATP, converting this energy to translocate unidirectionally, with different strand polarities and substrate binding specificities, along one strand of a nucleic acid. In doing so, DNA and RNA helicases separate duplex strands or remove nucleoprotein complexes, affecting DNA repair and the architecture of replication forks. In this review, we focus on recent advances on the roles and regulations of DNA helicases in homologous recombination repair, a critical pathway for mending damaged chromosomes and for ensuring genome integrity. - doi.org/10.1016/j.gde.2021.06.009 - 2021

PubReading [59] - Complexities of Viral Mutation Rates - K. Peck & A. Lauringa

Tue, 16 Nov 2021 21:00:21 GMT

Many viruses evolve rapidly. This is due, in part, to their high mutation rates. Mutation rate estimates for over 25 viruses are currently available. Here, we review the population genetics of virus mutation rates. We specifically cover the topics of mutation rate estimation, the forces that drive the evolution of mutation rates, and how the optimal mutation rate can be context-dependent. - doi.org/10.1128/JVI .01031-17. - 2018

PubReading [58] - Engineered miniature CRISPR-Cas system for mammalian genome regulation and editing - X. Xu, L. S. Qi et al.

Mon, 15 Nov 2021 21:00:20 GMT

Compact and versatile CRISPR-Cas systems will enable genome engineering applications through high-efficiency delivery in a wide variety of contexts. Here, we create an efficient miniature Cas system (CasMINI) engineered from the type V-F Cas12f (Cas14) system by guide RNA and protein engineering, which is less than half the size of currently used CRISPR systems (Cas9 or Cas12a). We demonstrate that CasMINI can drive high levels of gene activation (up to thousands-fold increases), while the natural Cas12f system fails to function in mammalian cells. We show that the CasMINI system has comparable activities to Cas12a for gene activation, is highly specific, and allows robust base editing and gene editing. We expect that CasMINI can be broadly useful for cell engineering and gene therapy applications ex vivo and in vivo. - doi.org/10.1016/j.molcel.2021.08.008 - 2021

PubReading [57] - Obesity and hypertension - G. Seravallea and G. Grassi

Tue, 09 Nov 2021 19:00:20 GMT

Obesity and in particular the excessive visceral fat distribution is accompanied by several alterations at hormonal, inflammatory and endothelial level. These alterations induce a stimulation of several other mechanisms that contribute to the hypertensive state and on the other side to increase the cardiovascular morbidity. In these chapter we will examine the main mechanisms of obesity and obesity-related hypertension and in particular the role of sympathetic nervous system, the alterations of the renal function and at the microvascular level. We will also depict the role of insulin resistance as factor stimulating and potentiating the other mechanisms. The second part will be focalized on the major target organ damage linked with obesity and obesity-related hypertension. We will finally describe the management and treatment of obesity and the antihypertensive drug therapies more effective in hypertensive obeses. - doi.org/10.1016/j.phrs.2017.05.013 - 2017

PubReading [56] - Hypertension and cardiovascular risk: General aspects - S. E. Kjeldsen

Mon, 08 Nov 2021 19:00:19 GMT

Hypertension is the strongest or one of the strongest risk factors for almost all different cardiovascular diseases acquired during life, including coronary disease, left ventricular hypertrophy and valvular heart diseases, cardiac arrhythmias including atrial fibrillation, cerebral stroke and renal failure. The continuous relationship between blood pressure and cardiovascular and renal events makes the distinction between high normal blood pressure and hypertension based on arbitrary cut-off values for blood pressures. Overall the prevalence of hypertension in different European countries appears to be around 30–45% of the general population, with a steep increase with ageing. The prevention of cardiovascular disease and treatment recommendations should be related to quantification of total cardiovascular risk which could be estimated from several different models. However the impact of age on risk is so strong that young adults (particularly women) are unlikely to reach high-risk levels even when they have more than one major risk factor and a clear increase in relative risk. Therefore age-adjusted models, models assessing relative risks compared to others of same age and models including thorough assessments of target organ damage and ambulatory 24 h blood pressure are needed together with national models because of the large variations between countries. - doi.org/10.1016/j.phrs.2017.11.003 - 2017

PubReading [55] - Protein phase separation and determinants of in cell crystallization - C. N. Mudogo, C. Betzel

Sat, 06 Nov 2021 19:00:20 GMT

Liquid-liquid phase separation (LLPS) in cells is known as a complex physicochemical process causing the formation of membrane-less organelles (MLOs). Cells have well-defined different membrane-surrounded organelles like mitochondria, endoplasmic reticulum, lysosomes, peroxisomes, etc., however, on demand they can create MLOs as stress granules, nucleoli and P bodies to cover vital functions and regulatory activities. However, the mechanism of intracellular molecule assembly into functional compartments within a living cell remains till now not fully understood. in vitro and in vivo investigations unveiled that MLOs emerge after preceding liquid-liquid, liquid gel, liquid-semi-crystalline, or liquid-crystalline phase separations. Liquid-liquid and liquid-gel MLOs form the majority of cellular phase separation events, while the occurrence of micro-sized crystals in cells was only rarely observed, however can be considered as a result of a preceding protein phase separation event. In vivo, also known and termed as in cellulo crystals, are reported since 1853. In some cases, they have been linked to vital cellular functions, such as storage and detoxification. How- ever, the occurrence of in cellulo crystals is also associated to diseases like cataract, hemoglobin C diseases, etc. Therefore, better knowledge about the involved molecular processes will support drug discovery investigations to cure diseases related to in cellulo crystallization. We summarize physical and chemical determinants known today required for phase separation initiation and formation and in cellulo crystal growth. In recent years it has been demonstrated that LLPS plays a crucial role in cell compartmentalization and formation of MLOs. Here we discuss potential mechanisms and potential crowding agents involved in protein phase separation and in cellulo crystallization. - DOI: 10.1111/tra.12711 - 2019

PubReading [54] - The Role of Inflammation in Diabetes: Current Concepts and Future Perspectives - S. Tsalamandris, D. Tousoulis et al

Fri, 05 Nov 2021 19:00:20 GMT

Diabetes is a complex metabolic disorder affecting the glucose status of the human body. Chronic hyperglycaemia related to diabetes is associated with end organ failure. The clinical relationship between diabetes and atherosclerotic cardiovascular disease is well established. This makes therapeutic approaches that simultaneously target diabetes and atherosclerotic disease an attractive area for research. The majority of people with diabetes fall into two broad pathogenetic categories, type 1 or type 2 diabetes. The role of obesity, adipose tissue, gut microbiota and pancreatic beta cell function in diabetes are under intensive scrutiny with several clinical trials to have been completed while more are in development. The emerging role of inflammation in both type 1 and type 2 diabetes (T1D and T1D) pathophysiology and associated metabolic disorders, has generated increasing interest in targeting inflammation to improve prevention and control of the disease. After an extensive review of the possible mechanisms that drive the metabolic pattern in T1D and T2D and the inflammatory pathways that are involved, it becomes ever clearer that future research should focus on a model of combined suppression for various inflammatory response pathways. - doi.org/10.15420/ecr.2018.33.1 - 2019

PubReading [53] - Molecular targeted therapy of glioblastoma - E. Le Rhun, M. Weller et al.

Thu, 04 Nov 2021 19:00:19 GMT

Glioblastomas are intrinsic brain tumors thought to originate from neuroglial stem or progenitor cells. More than 90% of glioblastomas are isocitrate dehydrogenase (IDH)-wildtype tumors. Incidence increases with age, males are more often affected. Beyond rare instances of genetic predisposition and irradiation exposure, there are no known glioblastoma risk factors. Surgery as safely feasible followed by involved-field radiotherapy plus concomitant and maintenance temozolomide chemotherapy define the standard of care since 2005. Except for prolonged progression-free, but not overall survival afforded by the vascular endothelial growth factor antibody, bevacizumab, no pharmacological intervention has been demonstrated to alter the course of disease. Specifically, targeting cellular pathways frequently altered in glioblastoma, such as the phosphoinositide 3- kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR), the p53 and the retinoblastoma (RB) pathways, or epidermal growth factor receptor (EGFR) gene amplification or mutation, have failed to improve outcome, likely because of redundant compensatory mechanisms, insufficient target coverage related in part to the blood brain barrier, or poor tolerability and safety. Yet, uncommon glioblastoma subsets may exhibit specific vulnerabilities amenable to targeted interventions, including, but not limited to: high tumor mutational burden, BRAF mutation, neurotrophic tryrosine receptor kinase (NTRK) or fibroblast growth factor receptor (FGFR) gene fusions, and MET gene amplification or fusions. There is increasing interest in targeting not only the tumor cells, but also the microenvironment, including blood vessels, the monocyte/macrophage/microglia compartment, or T cells. Improved clinical trial designs using pharmacodynamic endpoints in enriched patient populations will be required to develop better treatments for glioblastoma. - doi.org/10.1016/j.ctrv.2019.101896 - 2019

PubReading [52] - Choosing the Method of Crystallization to Obtain Optimal Results - L. Govada and N. E. Chayen

Wed, 03 Nov 2021 19:00:20 GMT

Anyone who has ever attempted to crystallise a protein or other biological macromolecule has encountered at least one, if not all of the following scenarios: No crystals at all, tiny low quality that do not diffract at all. In this paper, we review a number of simple ways to overcome such problems, which have worked well in our hands and in other laboratories. It brings together information that has been dispersed in various publications and lectures over the years and includes further information that has not been previously published. - doi:10.3390/cryst9020106 - 2019

PubReading [51] - p53, cancer and the immune response - J. Blagih, M. D. Buck and K. H. Vousden

Mon, 01 Nov 2021 21:00:21 GMT

The importance of cancer-cell-autonomous functions of the tumour suppressor p53 (encoded by TP53) has been established in many studies, but it is now clear that the p53 status of the cancer cell also has a profound impact on the immune response. Loss or mutation of p53 in cancers can affect the recruitment and activity of myeloid and T cells, allowing immune evasion and promoting cancer progression. p53 can also function in immune cells, resulting in various outcomes that can impede or support tumour development. Understanding the role of p53 in tumour and immune cells will help in the development of therapeutic approaches that can harness the differential p53 status of cancers compared with most normal tissue. - doi:10.1242/jcs.237453 - 2020

PubReading [50] - Ki-67 gene expression - S. Uxa, K. Engeland

Sat, 30 Oct 2021 20:00:24 GMT

Ki-67 serves as a prominent cancer marker. We describe how expression of the MKI67 gene coding for Ki-67 is controlled during the cell cycle. MKI67 mRNA and Ki-67 protein are maximally expressed in G2 phase and mitosis. Expression is dependent on two CHR elements and one CDE site in the MKI67 promoter. DREAM transcriptional repressor complexes bind to both CHR sites and downregulate the expression in G0/G1 cells. Upregulation of MKI67 transcription coincides with binding of B-MYB-MuvB and FOXM1-MuvB complexes from S phase into G2/M. Importantly, binding of B-MYB to the two CHR elements correlates with loss of CHR-dependent MKI67 promoter activation in B-MYB-knockdown experiments. In knockout cell models, we find that DREAM/MuvB-dependent transcriptional control cooperates with the RB Retinoblastoma tumor suppressor. Furthermore, the p53 tumor suppressor indirectly downregulates transcription of the MKI67 gene. This repression by p53 requires p21/CDKN1A. These results are consistent with a model in which DREAM, B-MYB-MuvB, and FOXM1-MuvB together with RB cooperate in cell cycle-dependent transcription and in transcriptional repression following p53 activation. In conclusion, we present mechanisms how MKI67 gene expression followed by Ki-67 protein synthesis is controlled during the cell cycle and upon induction of DNA damage, as well as upon p53 activation. - doi.org/10.1038/s41418-021-00823-x - 2021

PubReading [49] - Rationally designed ruthenium complexes for 1- and 2-photon photodynamic therapy - J. Karges, G. Gasser et al.

Fri, 29 Oct 2021 20:00:24 GMT

The use of photodynamic therapy (PDT) against cancer has received increasing attention over recent years. However, the application of the currently approved photosensitizers (PSs) is limited by their poor aqueous solubility, aggregation, photobleaching and slow clearance from the body. To overcome these limitations, there is a need for the development of new classes of PSs with ruthenium(II) polypyridine complexes currently gaining momentum. However, these compounds generally lack significant absorption in the biological spectral window, limiting their application to treat deep-seated or large tumors. To overcome this drawback, ruthenium(II) polypyridine complexes designed in silico with (E,E′)-4,4′-bisstyryl- 2,2′-bipyridine ligands show impressive 1- and 2-Photon absorption up to a magnitude higher than the ones published so far. While nontoxic in the dark, these compounds are phototoxic in various 2D monolayer cells, 3D multicellular tumor spheroids and are able to eradicate a multiresistant tumor inside a mouse model upon clinically relevant 1-Photon and 2-Photon excitation. - doi.org/10.1038/s41467-020-16993-0 - 2020

PubReading [48] - Induction of ubiquitin C (UBC) gene transcription is mediated by HSF1: role of proteotoxic and oxidative stress - M. Bianchi, M. Magnani et al.

Thu, 28 Oct 2021 20:00:25 GMT

The polyubiquitin gene ubiquitin C (UBC) is considered a stress protective gene and is upregulated under various stressful conditions, which is probably a consequence of an increased demand for ubiquitin in order to remove toxic misfolded proteins. We previously identified heat shock elements (HSEs) within the UBC promoter, which are responsible for heat shock factor (HSF)1-driven induction of the UBC gene and are activated by proteotoxic stress. Here, we determined the molecular players driving the UBC gene transcriptional response to arsenite treatment, mainly addressing the role of the nuclear factor-erythroid 2-related factor 2 (Nrf2)-mediated antioxidant pathway. Exposure of HeLa cells to arsenite caused a time-dependent increase of UBC mRNA, while cell viability and proteasome activity were not affected. Nuclear accumulation of HSF1 and Nrf2 transcription factors were detected upon both arsenite and MG132 treatment, while HSF2 nuclear levels increased in MG132-treated cells. Notably, siRNA-mediated knockdown of Nrf2 did not reduce UBC transcription under either basal or stressful conditions, but significantly impaired the constitutive and inducible expression of well-known antioxidant response element-dependent genes. A chromatin immunoprecipitation assay consistently failed to detect Nrf2 binding to the UBC promoter sequence. By contrast, depletion of HSF1, but not HSF2, significantly compromised stress-induced UBC expression. Critically, HSF1-mediated UBC trans-activation upon arsenite exposure relies on transcription factor binding to previously mapped distal HSEs, as demonstrated to occur under proteasome inhibition. These data highlight HSF1 as the pivotal transcription factor that translates different stress signals into UBC gene transcriptional induction. - doi:10.1002/2211-5463.12484 - 2018

PubReading [46] - DNA-mediated engineering of multicomponent enzyme crystals - J. D. Brodina, C. A. Mirkin et al.

Tue, 26 Oct 2021 20:00:20 GMT

The ability to predictably control the coassembly of multiple nanoscale building blocks, especially those with disparate chemical and physical properties such as biomolecules and inorganic nanoparticles, has far-reaching implications in catalysis, sensing, and photonics, but a generalizable strategy for engineering specific contacts between these particles is an outstanding challenge. This is especially true in the case of proteins, where the types of possible interparticle interactions are numerous, diverse, and complex. Herein, we explore the concept of trading protein–protein interactions for DNA–DNA interactions to direct the assembly of two nucleic-acid–functionalized proteins with distinct surface chemstries into six unique lattices composed of catalytically active proteins, or of a combination of proteins and DNA-modified gold nanoparticles. The programmable nature of DNA–DNA interactions used in this strategy allows us to control the lattice symmetries and unit cell constants, as well as the compositions and habit, of the resulting crystals. This study provides a potentially generalizable strategy for constructing a unique class of materials that take advantage of the diverse morphologies, surface chemistries, and functionalities of proteins for assembling functional crystalline materials. - doi/10.1073/pnas.1503533112 - 2015

PubReading [45] - Built-In Active Microneedle Patch with Enhanced Autonomous Drug Delivery - M.A. Lopez-Ramirez, J. Wang et al.

Mon, 25 Oct 2021 20:00:20 GMT

The use of microneedles has facilitated the painless localized delivery of drugs across the skin. However, their efficacy has been limited by slow diffusion of molecules and often requires external triggers. Herein, an autonomous and degradable, active microneedle delivery platform is introduced, employing magnesium microparticles loaded within the microneedle patch, as the built-in engine for deeper and faster intradermal payload delivery. The magnesium particles react with the interstitial fluid, leading to an explosive-like rapid production of H2 bubbles, providing the necessary force to breach dermal barriers and enhance payload delivery. The release kinetics of active microneedles is evaluated in vitro by measuring the amount of IgG antibody (as a model drug) that passed through phantom tissue and a pigskin barrier. In vivo experiments using a B16F10 mouse melanoma model demonstrate that the active delivery of anti-CTLA-4 (a checkpoint inhibitor drug) results in greatly enhanced immune response and significantly longer survival. Moreover, spatially resolved zones of active and passive microneedles allow a combinatorial rapid burst response along with slow, sustained release, respectively. Such versatile and effective autonomous dynamic microneedle delivery technology offers considerable promise for a wide range of therapeutic applications, toward a greatly enhanced outcome, convenience, and cost. - DOI: 10.1002/adma.201905740 - 2020

PubReading [44] - I-motif DNA structures are formed in the nuclei of human cells - M. Zeraati, D. Christ et al.

Sat, 23 Oct 2021 20:00:20 GMT

Human genome function is underpinned by the primary storage of genetic information in canonical B-form DNA, with a second layer of DNA structure providing regulatory control. I-motif structures are thought to form in cytosine-rich regions of the genome and to have regulatory functions; however, in vivo evidence for the existence of such structures has so far remained elusive. Here we report the generation and characterization of an antibody fragment (iMab) that recognizes i-motif structures with high selectivity and affinity, enabling the detection of i-motifs in the nuclei of human cells. We demonstrate that the in vivo formation of such structures is cell-cycle and pH dependent. Furthermore, we provide evidence that i-motif structures are formed in regulatory regions of the human genome, including promoters and telomeric regions. Our results support the notion that i-motif structures provide key regulatory roles in the genome. - doi.org/10.1038/s41557-018-0046-3 - 2018

PubReading [43] - Click and Cut: a click chemistry approach to developing oxidative DNA damaging agents - N. McStay, A. Kellett et al.

Fri, 22 Oct 2021 20:00:20 GMT

Metallodrugs provide important first-line treatment against various forms of human cancer. To overcome chemotherapeutic resistance and widen treatment possibilities, new agents with improved or alternative modes of action are highly sought after. Here, we present a click chemistry strategy for developing DNA damaging metallodrugs. The approach in- volves the development of a series of polyamine ligands where three primary, secondary or tertiary alkyne-amines were selected and ‘clicked’ using the copper-catalysed azide-alkyne cycloaddition reac- tion to a 1,3,5-azide mesitylene core to produce a family of compounds we call the ‘Tri-Click’ (TC) se- ries. From the isolated library, one dominant ligand (TC1) emerged as a high-affinity copper(II) binding agent with potent DNA recognition and damaging properties. Using a range of in vitro biophysical and molecular techniques––including free radical scavengers, spin trapping antioxidants and base excision repair (BER) enzymes–the oxidative DNA damaging mechanism of copper-bound TC1 was elucidated. This activity was then compared to intracellular results obtained from peripheral blood mononuclear cells exposed to Cu(II)–TC1 where use of BER enzymes and fluorescently modified dNTPs enabled the characterisation and quantification of genomic DNA lesions produced by the complex. The approach can serve as a new avenue for the design of DNA damaging agents with unique activity profiles. - doi.org/10.1093/nar/gkab817 - 2021

PubReading [42] - Circulating tumoral DNA: Preanalytical validation and quality control in a diagnostic laboratory - S. Nikolaev, T. Nouspikel et al.

Wed, 20 Oct 2021 23:00:20 GMT

We present the results of our technical validation process in establishing the analysis of circulating tumor DNA (ctDNA) as a diagnostic tool. Like most cells in our body, tumor cells shed DNA in the blood flow. Analysis of ctDNA mutational content can provide invaluable information on the genetic makeup of a tumor, and assist oncologists in deciding on therapy, or in following residual disease. However, low absolute amounts of circulating DNA and low tumor fraction constitute formidable analytical challenges. A key step is to avoid contamination with genomic DNA from cell lysis. Several brands of specialized blood collection tubes are available to prevent leukocyte lysis. We show that they are not equally efficient, depending on storage temperature and time before plasma preparation. We report our analysis of preanalytical factors pertaining to ctDNA analysis (tubes, transportation time, temperature) and our conclusions in terms of instructions to prescribing physicians. We also stress the importance of proper DNA quality control and compare several methods, including a differential amplicon length PCR technique which allows determination of multiple QC parameters from minimal amounts of DNA. Altogether, these data provide useful practical information to diagnostic laboratories wishing to implement the assay of ctDNA in clinical practice. - doi.org/10.1016/j.ab.2017.11.004 - 2018

PubReading [41] - Apoptosis: A Target for Anticancer Therapy - C. M. Pfeffer and A. T. K. Singh

Wed, 20 Oct 2021 20:00:20 GMT

Apoptosis, the cell’s natural mechanism for death, is a promising target for anticancer therapy. Both the intrinsic and extrinsic pathways use caspases to carry out apoptosis through the cleavage of hundreds of proteins. In cancer, the apoptotic pathway is typically inhibited through a wide variety of means including overexpression of antiapoptotic proteins and under-expression of proapoptotic proteins. Many of these changes cause intrinsic resistance to the most common anticancer therapy, chemotherapy. Promising new anticancer therapies are plant-derived compounds that exhibit anticancer activity through activating the apoptotic pathway. doi:10.3390/ijms19020448 - 2018

PubReading [40] - Phosphoproteome profiling uncovers a key role for CDKs in TNF signaling - M. Tanzer, M. Mann et al

Tue, 19 Oct 2021 20:00:19 GMT

Tumor necrosis factor (TNF) is one of the few cytokines successfully targeted by therapies against inflammatory diseases. However, blocking this well studied and pleiotropic ligand can cause dramatic side-effects. Here, we reason that a systems-level proteomic analysis of TNF signaling could dissect its diverse functions and offer a base for developing more targeted therapies. Therefore, we combine phosphoproteomics time course experiments with sub-cellular localization and kinase inhibitor analysis to identify functional modules of protein phosphorylation. The majority of regulated phosphorylation events can be assigned to an upstream kinase by inhibiting master kinases. Spatial proteomics reveals phosphorylation- dependent translocations of hundreds of proteins upon TNF stimulation. Phosphoproteome analysis of TNF-induced apoptosis and necroptosis uncovers a key role for transcriptional cyclin-dependent kinase activity to promote cytokine production and prevent excessive cell death downstream of the TNF signaling receptor. This resource of TNF-induced pathways and sites can be explored at http://tnfviewer.biochem.mpg.de/. - doi.org/10.1038/s41467-021-26289-6 - 2021

PubReading [39] - Crystal structure of a membrane-bound metalloenzyme that catalyses the biological oxidation of methane - R. L. Lieberman and A. C. Rosenzweig

Mon, 18 Oct 2021 22:14:18 GMT

Particulate methane monooxygenase (pMMO) is an integral membrane metalloenzyme that catalyses the conversion of methane to methanol. Knowledge of how pMMO performs this extremely challenging chemistry may have an impact on the use of methane as an alternative energy source by facilitating the development of new synthetic catalysts. We have determined the structure of pMMO from the methanotroph Methylococcus capsulatus (Bath) to a resolution of 2.8A ̊. The enzyme is a trimer with an a3b3g3 polypeptide arrangement. Two metal centres, modelled as mononuclear copper and dinuclear copper, are located in soluble regions of each pmoB subunit, which resembles cytochrome c oxidase subunit II. A third metal centre, occupied by zinc in the crystal, is located within the membrane. The structure provides new insight into the molecular details of biological methane oxidation. - doi.org/10.1038/nature03311 - 2005

PubReading [38] - PeptiCHIP: A Microfluidic Platform for Tumor Antigen Landscape Identification - S. Feola, V. Cerullo et al.

Sat, 16 Oct 2021 20:00:21 GMT

Identification of HLA class I ligands from the tumor surface (ligandome or immunopeptidome) is essential for designing T-cell mediated cancer therapeutic approaches. However, the sensitivity of the process for isolating MHC-I restricted tumor-specific peptides has been the major limiting factor for reliable tumor antigen characterization, making clear the need for technical improvement. Here, we describe our work from the fabrication and development of a microfluidic-based chip (PeptiCHIP) and its use to identify and characterize tumor-specific ligands on clinically relevant human samples. Specifically, we assessed the potential of immobilizing a pan-HLA antibody on solid surfaces via well-characterized streptavidin−biotin chemistry, overcoming the limitations of the cross-linking chemistry used to prepare the affinity matrix with the desired antibodies in the immunopeptidomics workflow. Furthermore, to address the restrictions related to the handling and the limited availability of tumor samples, we further developed the concept toward the implementation of a microfluidic through-flow system. Thus, the biotinylated pan-HLA antibody was immobilized on streptavidin-functionalized surfaces, and immune-affinity purification (IP) was carried out on customized microfluidic pillar arrays made of thiol−ene polymer. Compared to the standard methods reported in the field, our methodology reduces the amount of antibody and the time required for peptide isolation. In this work, we carefully examined the specificity and robustness of our customized technology for immunopeptidomics workflows. We tested this platform by immunopurifying HLA-I complexes from 1 × 106 cells both in a widely studied B-cell line and in patients-derived ex vivo cell cultures, instead of 5 × 108 cells as required in the current technology. After the final elution in mild acid, HLA-I-presented peptides were identified by tandem mass spectrometry and further investigated by in vitro methods. These results highlight the potential to exploit microfluidics-based strategies in immunopeptidomics platforms and in personalized immunopeptidome analysis from cells isolated from individual tumor biopsies to design tailored cancer therapeutic vaccines. Moreover, the possibility to integrate multiple identical units on a single chip further improves the throughput and multiplexing of these assays with a view to clinical needs. - doi.org/10.1021/acsnano.1c04371 - 2021

PubReading [37] - High resolution single particle cryo-electron microscopy using beam-image shift - A. Cheng, B. Carragher et al.

Fri, 15 Oct 2021 20:00:21 GMT

Automated data acquisition is used widely for single-particle reconstruction of three-dimensional (3D) volumes of biological complexes preserved in vitreous ice and imaged in a transmission electron microscope. Automation has become integral to this method because of the very large number of particle images required in order to overcome the typically low signal-to-noise ratio of these images. For optimal efficiency, automated data acquisition software packages typically employ some beam-image shift targeting as this method is both fast and accurate ( ± 0.1 μm). In contrast, using only stage movement, relocation to a targeted area under low-dose conditions can only be achieved in combination with multiple iterations or long relaxation times, both reducing efficiency. Nevertheless it is well known that applying beam- image shift induces beam-tilt and with it a potential structure phase error with a phase error π/4 the highest acceptable value. This theory has been used as an argument against beam-image shift for high resolution data collection. Nevertheless, in practice many small beam-image shift datasets have resulted in 3D reconstructions beyond the π/4 phase error limit. To address this apparent contradiction, we performed cryo-EM single-particle reconstructions on a T20S proteasome sample using applied beam-image shifts corresponding to beam tilts from 0 to 10 mrad. To evaluate the results we compared the FSC values, and examined the water density peaks in the 3D map. We conclude that the phase error does not limit the validity of the 3D reconstruction from single-particle averaging beyond the π/4 resolution limit. - doi.org/10.1016/j.jsb.2018.07.015 - 2018

PubReading [36] - APE1: A skilled nucleic acid surgeon - A. Whitaker and B. Freudenthal

Thu, 14 Oct 2021 20:00:21 GMT

Before a deleterious DNA lesion can be replaced with its undamaged counterpart, the lesion must first be removed from the genome. This process of removing and replacing DNA lesions is accomplished by the careful coordination of several protein factors during DNA repair. One such factor is the multifunctional enzyme human apurinic/apyrimidinic endonuclease 1 (APE1), known best for its DNA backbone cleavage activity at AP sites during base excision repair (BER). APE1 preforms AP site incision with surgical precision and skill, by sculpting the DNA to place the cleavage site in an optimal position for nucleophilic attack within its compact protein active site. APE1, however, has demonstrated broad surgical expertise, and applies its DNA cleavage activity to a wide variety of DNA and RNA substrates. Here, we discuss what is known and unknown about APE1 cleavage mechanisms, focusing on structural and mechanistic considerations. Importantly, disruptions in the biological functions associated with APE1 are linked to numerous human maladies, including cancer and neurodegenerative diseases. The continued elucidation of APE1 mechanisms is required for rational drug design towards novel and strategic ways to target its associated repair pathways. - doi.org/10.1016/j.dnarep.2018.08.012 - 2018

PubReading [35] - Oligonucleotide conjugated multi-functional adeno-associated viruses - D. Katrekar, P. Mali et al.

Wed, 13 Oct 2021 20:00:21 GMT

Recombinant adeno-associated viruses (AAVs) are among the most commonly used vehicles for in vivo gene delivery. However, their tropism is limited, and additionally their efficacy can be negatively affected by prevalence of neutralizing antibodies in sera. Methodologies to systematically engineer AAV capsid properties would thus be of great relevance. In this regard, we develop here multi-functional AAVs by engineering precision tethering of oligonucleotides onto the AAV surface, and thereby enabling a spectrum of nucleic-acid programmable functionalities. Towards this, we engineered genetically encoded incorporation of unnatural amino acids (UAA) bearing bio-orthogonal chemical handles onto capsid proteins. Via these we enabled site-specific coupling of oligonucleotides onto the AAV capsid surface using facile click chemistry. The resulting oligo-AAVs could be sequence specifically labeled, and also patterned in 2D using DNA array substrates. Additionally, we utilized these oligo conjugations to engineer viral shielding by lipid-based cloaks that efficaciously protected the AAV particles from neutralizing serum. We confirmed these ‘cloaked AAVs’ retained full functionality via their ability to transduce a range of cell types, and also enable robust delivery of CRISPR-Cas9 effectors. Taken together, we anticipate this programmable oligo-AAV system will have broad utility in synthetic biology and AAV engineering applications. - DOI:10.1038/s41598-018-21742-x - 2018

PubReading [337] - Training early career researchers to use meta- research to improve science- A participant- guided “learn by doing” approach - T. Weissgerber

Mon, 12 Jun 2023 11:00:14 GMT

Meta-research, or the science of science, is a powerful technique that scientists can use to improve science, however most scientists are unaware that meta-research exists and courses are rare. This initiative demonstrates the feasibility of a participant-guided “learn by doing” approach, in which a multidisciplinary, global team of early career researchers learned meta-research skills by working together to design, conduct and publish a meta-research study.https://doi.org/10.1371/journal.pbio.3001073 - 2021

PubReading [34] - Replication protein A binds RNA and promotes R-Loop formation - O. Mazina, A. V. Mazin et al.

Tue, 12 Oct 2021 20:00:20 GMT

Replication protein A (RPA), a major eukaryotic ssDNA- binding protein, is essential for all metabolic processes that involve ssDNA, including DNA replication, repair, and damage signaling. To perform its functions, RPA binds ssDNA tightly. In contrast, it was presumed that RPA binds RNA weakly. However, recent data suggest that RPA may play a role in RNA metabolism. RPA stimulates RNA-templated DNA repair in vitro and associates in vivo with R-loops, the three-stranded structures consisting of an RNA-DNA hybrid and the displaced ssDNA strand. R-loops are common in the genomes of pro- and eukaryotes, including humans, and may play an important role in transcription-coupled homologous recombination and DNA replication restart. However, the mechanism of R-loop formation remains unknown. Here, we investigated the RNA-binding properties of human RPA and its possible role in R-loop formation. Using gel-retardation and RNA/ DNA competition assays, we found that RPA binds RNA with an unexpectedly high affinity (KD ’ 100 pM). Furthermore, RPA, by forming a complex with RNA, can promote R-loop formation with homologous dsDNA. In reconstitution experiments, we showed that human DNA polymerases can utilize RPA-generated R-loops for initiation of DNA synthesis, mimicking the process of replication restart in vivo. These results demonstrate that RPA binds RNA with high affinity, supporting the role of this protein in RNA metabolism and suggesting a mechanism of genome maintenance that depends on RPA- mediated DNA replication restart. - DOI 10.1074/jbc.RA120.013812 - 2020

PubReading [33] - Oxidative DNA Cleavage with Clip-Phenanthroline Triplex-Forming Oligonucleotide Hybrids - A. Panattoni, M. Hocek et al.

Mon, 11 Oct 2021 20:00:22 GMT

A systematic study of several new types of hybrids of Cu-chelated clamped phenanthroline artificial metallonuclease (AMN) with triplex-forming oligonucleotides (TFO) for sequence-specific cleavage of double-stranded DNA (dsDNA) is reported. The synthesis of these AMN–TFO hybrids is based on application of the alkyne–azide cycloaddition click reaction as the key step. The AMN was attached through different linkers at either the 5’- or 3’-ends or in the middle of the TFO stretch. The diverse hybrids efficiently formed triplexes with the target purine-rich sequence and their copper complexes were studied for their ability to cleave dsDNA in the presence of ascorbate as a reductant. In all cases, the influence of the nature and length of the AMN–TFO, time, conditions and amounts of ascorbate were studied, and optimum conjugates and a procedure that gave reasonably efficient (up to 34%) cleavage of the target sequence, while rendering an off-target dsDNA intact, were found. The footprint of cleavage on PAGE was identified only in one case, with low conversion; this means that cleavage does not proceed with single nucleotide precision. On the other hand, these AMN–TFO hybrids are useful for the selective degradation of target dsDNA sequences. Future improvements to this design may provide higher resolution and selectivity. - doi.org/10.1002/cbic.201900670. - 2019

PubReading [32] - Enhancing Protein Crystallization under a Magnetic Field - S. Young Ryu, H. Kyu Song et al.

Fri, 08 Oct 2021 22:17:49 GMT

High-quality crystals are essential to ensure high-resolution structural information. Protein crystals are controlled by many factors, such as pH, temperature, and the ion concentration of crystalline solutions. We previously reported the development of a device dedicated to protein crystallization. In the current study, we have further modified and improved our device. Exposure to external magnetic field leads to alignment of the crystal toward a preferred direction depending on the magnetization energy. Each material has different magnetic susceptibilities depending on the individual direction of their unit crystal cells. One of the strategies to acquire a large crystal entails controlling the nucleation rate. Furthermore, exposure of a crystal to a magnetic field may lead to new morphologies by affecting the crystal volume, shape, and quality. - doi:10.3390/cryst10090821 - 2020

PubReading [31] - TLR8 Is a Sensor of RNase T2 Degradation Products - W. Greulic, T. Carell, V. Hornung et al.

Thu, 07 Oct 2021 22:37:50 GMT

TLR8 is among the highest-expressed pattern-recognition receptors in the human myeloid compartment, yet its mode of action is poorly understood. TLR8 engages two distinct ligand binding sites to sense RNA degradation products, although it remains unclear how these ligands are formed in cellulo in the context of complex RNA molecule sensing. Here, we identified the lysosomal endoribonuclease RNase T2 as a non-redundant upstream component of TLR8- dependent RNA recognition. RNase T2 activity is required for rendering complex single-stranded, exogenous RNA molecules detectable for TLR8. This is due to RNase T2’s preferential cleavage of single-stranded RNA molecules between purine and uridine residues, which critically contributes to the supply of catabolic uridine and the generation of purine-2',3'-cyclophosphate-terminated oligoribonucleotides. Thus-generated molecules constitute agonistic ligands for the first and second binding pocket of TLR8. Together, these results establish the identity and origin of the RNA-derived molecular pattern sensed by TLR8. - doi.org/10.1016/j.cell.2019.11.001 - 2019

PubReading [30] - Cone-shaped HIV-1 capsids are transported through intact nuclear pores - V. Zila, M. Beck et al.

Wed, 06 Oct 2021 20:00:40 GMT

Human immunodeficiency virus (HIV-1) remains a major health threat. Viral capsid uncoating and nuclear import of the viral genome are critical for productive infection. The size of the HIV-1 capsid is generally believed to exceed the diameter of the nuclear pore complex (NPC), indicating that capsid uncoating has to occur prior to nuclear import. Here, we combined correlative light and electron microscopy with subtomogram averaging to capture the structural status of reverse transcription-competent HIV-1 complexes in infected T cells. We demonstrated that the diameter of the NPC in cellulo is sufficient for the import of apparently intact, cone-shaped capsids. Subsequent to nuclear import, we detected disrupted and empty capsid fragments, indicating that uncoating of the replication complex occurs by breaking the capsid open, and not by disassembly into individual subunits. Our data directly visualize a key step in HIV-1 replication and enhance our mechanistic understanding of the viral life cycle. - doi.org/10.1016/j.cell.2021.01.025 - 2021

PubReading [29] - Using antibodies to control DNA-templated chemical reactions - L. B. Pellejero, F. Ricci, T. Brown Jr et al.

Tue, 05 Oct 2021 20:00:19 GMT

DNA-templated synthesis takes advantage of the programmability of DNA-DNA interactions to accelerate chemical reactions under diluted conditions upon sequence-specific hybridization. While this strategy has proven advantageous for a variety of applications, including sensing and drug discovery, it has been so far limited to the use of nucleic acids as templating elements. Here, we report the rational design of DNA templated synthesis controlled by specific IgG antibodies. Our approach is based on the co-localization of reactants induced by the bivalent binding of a specific IgG antibody to two antigen-conjugated DNA templating strands that triggers a chemical reaction that would be otherwise too slow under diluted conditions. This strategy is versatile, orthogonal and adaptable to different IgG antibodies and can be employed to achieve the targeted synthesis of clinically-relevant molecules in the presence of specific IgG biomarker antibodies. - doi.org/10.1038/s41467-020-20024-3 - 2020

PubReading [28] - DNA G‐quadruplexes in the human genome: detection, functions and therapeutic potential - R. Hänsel-Hertsch, M. Di Antonio and S. Balasubramanian

Mon, 04 Oct 2021 20:00:19 GMT

Single-stranded guanine-rich DNA sequences can fold into four-stranded DNA structures called G-quadruplexes (G4s) that arise from the self-stacking of two or more guanine quartets. There has been considerable recent progress in the detection and mapping of G4 structures in the human genome and in biologically relevant contexts. These advancements, many of which align with predictions made previously in computational studies, provide important new insights into the functions of G4 structures in, for example, the regulation of transcription and genome stability, and uncover their potential relevance for cancer therapy. - doi:10.1038/nrm.2017.3 - 2017

PubReading [27] - Rapid Generation of Long Noncoding RNA Knockout Mice Using CRISPR/Cas9 Technology - N. R. Hansmeier, J.W. Kornfeld et al.

Fri, 01 Oct 2021 22:29:21 GMT

In recent years, long noncoding RNAs (lncRNAs) have emerged as multifaceted regulators of gene expression, controlling key developmental and disease pathogenesis processes. However, due to the paucity of lncRNA loss-of-function mouse models, key questions regarding the involvement of lncRNAs in organism homeostasis and (patho)-physiology remain difficult to address experimentally in vivo. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 platform provides a powerful genome-editing tool and has been successfully applied across model organisms to facilitate targeted genetic mutations, including Caenorhabditis elegans, Drosophila melanogaster, Danio rerio and Mus musculus. However, just a few lncRNA-deficient mouse lines have been created using CRISPR/Cas9-mediated genome engineering, presumably due to the need for lncRNA-specific gene targeting strategies considering the absence of open-reading frames in these loci. Here, we describe a step-wise procedure for the generation and validation of lncRNA loss-of-function mouse models using CRISPR/Cas9-mediated genome engineering. In a proof-of-principle approach, we generated mice deficient for the liver-enriched lncRNA Gm15441, which we found downregulated during development of metabolic disease and induced during the feeding/fasting transition. Further, we discuss guidelines for the selection of lncRNA targets and provide protocols for in vitro single guide RNA (sgRNA) validation, assessment of in vivo gene-targeting efficiency and knockout confirmation. The procedure from target selection to validation of lncRNA knockout mouse lines can be completed in 18–20 weeks, of which

PubReading [26] - Tools and methods for circular dichroism spectroscopy of proteins: a tutorial review - A. J. Miles, Robert W. Janes and B. A. Wallace

Wed, 29 Sep 2021 21:00:19 GMT

Circular dichroism (CD) spectroscopy is a widely-used method in biochemistry, structural biology and pharmaceutical chemistry. More than 24000 papers published in the past decade have included CD characterisations of proteins; many of those studies have also included other complementary chemical, biophysical, and computational chemistry methods. This tutorial review describes the background to the technique of CD spectroscopy and good practice methods for high quality data collection. It specifically focuses on both established and new methods and tools available for experimental design and interpretation, data processing, visualisation, analysis, validation, archiving, and accession, including tools developed to enhance the complementarity of this method with other structural and chemical biology studies. - DOI: 10.1039/d0cs00558d - 2021

PubReading [25] - Brown adipose tissue monocytes support tissue expansion - A. Gallerand, L. Yvan-Charvet1 & S. Ivanov et al.

Tue, 28 Sep 2021 21:00:19 GMT

Monocytes are part of the mononuclear phagocytic system. Monocytes play a central role during inflammatory conditions and a better understanding of their dynamics might open therapeutic opportunities. In the present study, we focused on the characterization and impact of monocytes on brown adipose tissue (BAT) functions during tissue remodeling. Single-cell RNA sequencing analysis of BAT immune cells uncovered a large diversity in monocyte and macrophage populations. Fate-mapping experiments demonstrated that the BAT macrophage pool requires constant replenishment from monocytes. Using a genetic model of BAT expansion, we found that brown fat monocyte numbers were selectively increased in this scenario. This observation was confirmed using a CCR2-binding radiotracer and positron emission tomography. Importantly, in line with their tissue recruitment, blood monocyte counts were decreased while bone marrow hematopoiesis was not affected. Monocyte depletion prevented brown adipose tissue expansion and altered its architecture. Podoplanin engagement is strictly required for BAT expansion. Together, these data redefine the diversity of immune cells in the BAT and emphasize the role of monocyte recruitment for tissue remodeling. - doi.org/10.1038/s41467-021-25616-1 - 2021

PubReading [24] - Root of the Tree: The Significance, Evolution, and Origins of the Ribosome - J. C. Bowman, L. D. Williams et al.

Mon, 27 Sep 2021 21:00:19 GMT

The ribosome is an ancient molecular fossil that provides a telescope to the origins of life. Made from RNA and protein, the ribosome translates mRNA to coded protein in all living systems. Universality, economy, centrality and antiquity are ingrained in translation. The translation machinery dominates the set of genes that are shared as orthologues across the tree of life. The lineage of the translation system defines the universal tree of life. The function of a ribosome is to build ribosomes; to accomplish this task, ribosomes make ribosomal proteins, polymerases, enzymes, and signaling proteins. Every coded protein ever produced by life on Earth has passed through the exit tunnel, which is the birth canal of biology. During the root phase of the tree of life, before the last common ancestor of life (LUCA), exit tunnel evolution is dominant and unremitting. Protein folding coevolved with evolution of the exit tunnel. The ribosome shows that protein folding initiated with intrinsic disorder, supported through a short, primitive exit tunnel. Folding progressed to thermodynamically stable β- structures and then to kinetically trapped α-structures. The latter were enabled by a long, mature exit tunnel that partially offset the general thermodynamic tendency of all polypeptides to form β-sheets. RNA chaperoned the evolution of protein folding from the very beginning. The universal common core of the ribosome, with a mass of nearly 2 million Daltons, was finalized by LUCA. The ribosome entered stasis after LUCA and remained in that state for billions of years. Bacterial ribosomes never left stasis. Archaeal ribosomes have remained near stasis, except for the superphylum Asgard, which has accreted rRNA post LUCA. Eukaryotic ribosomes in some lineages appear to be logarithmically accreting rRNA over the last billion years. Ribosomal expansion in Asgard and Eukarya has been incremental and iterative, without substantial remodeling of pre-existing basal structures. The ribosome preserves information on its history. - dx.doi.org/10.1021/acs.chemrev.9b00742 - 2020

PubReading [23] - Deformylation of 5-Formylcytidine in Different Cell Types - E. Korytiakovà and T. Carell

Thu, 09 Sep 2021 21:41:08 GMT

Epigenetic programming of cells requires methylation of deoxycytidines (dC) to 5-methyl-dC (mdC) followed by oxidation to 5-hydroxymethyl-dC (hmdC), 5-formyl-dC (fdC), and 5-carboxy-dC (cadC). Subsequent transformation of fdC and cadC back to dC by various pathways establishes a chemical intra-genetic control circle. One of the discussed pathways involves the Tdg-independent deformylation of fdC directly to dC. Here we report the synthesis of a fluorinated fdC feeding probe (F-fdC) to study direct deformylation to F-dC. The synthesis was performed along a novel pathway that circumvents any F-dC as a reaction intermediate to avoid contamination interference. Feeding of F-fdC and observation of F-dC formation in vivo allowed us to gain insights into the Tdg-independent removal process. While deformylation was shown to occur in stem cells, we here provide data that prove deformylation also in different somatic cell types. We also investigated active demethylation in a non-dividing neurogenin-inducible system of iPS cells that differentiate into bipolar neurons. - doi.org/10.1002/anie.202107089. - 2021

PubReading [22] - Shining Light on CRISPR Gene Editing - L. Taemaitree and T. Brown

Tue, 07 Sep 2021 21:09:49 GMT

Advanced spatiotemporal control of CRISPR-Cas9 activity is demonstrated through the use of chemically modified photoactivatable guide RNA. - doi.org/10.1021/acscentsci.0c00350 - 2020

PubReading [21] - DNA structure from A to B - R.Dickerson and HL. Ng

Mon, 06 Sep 2021 22:13:40 GMT

P. Shing Ho and his colleagues at Oregon State and Berkeley publish in this issue of PNAS an interesting study (1) of helical structure in the DNA hexamer GGCGCC, finding that various states that appear to be logical intermediates between A-DNA and B-DNA can be induced by methylation or bromination of cytosine or by crystal packing. Their results bear on three issues that have been argued over in the past: (i) the differences between A-DNA and B- DNA and transitions between them, (ii) the intrinsic sequence-dependent malleability of a DNA duplex, and (iii) the effects of local helix packing on DNA fine-structure. - doi-10.1073-pnas.141238898 - 2001

PubReading [18] - Controlling and enhancing CRISPR systems - H. Shivram, B. Cress, G. Knott, J. Doudna

Wed, 01 Sep 2021 22:53:02 GMT

Many bacterial and archaeal organisms use CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR associated) systems to defend themselves from mobile genetic elements. These CRISPR-Cas systems are classified into six types based on their composition and mechanism. CRISPR-Cas enzymes are widely used for genome editing and offer immense therapeutic opportunity to treat genetic diseases. To realize their full potential, it is important to control the timing, duration, efficiency and specificity of CRISPR-Cas enzyme activities. In this review we discuss the mechanisms of natural CRISPR-Cas regulatory biomolecules that enhance or inhibit CRISPR-Cas immunity by altering enzyme function. We also discuss the potential applications of these CRISPR regulators and highlight unanswered questions about their evolution and purpose in nature. - doi:10.1038/s41589-020-00700-7 - 2021

PubReading [17] - Antisense, RNAi, and gene silencing strategies for therapy: Mission possible or impossible? - E. Rayburn and R. Zhang

Tue, 31 Aug 2021 22:24:54 GMT

Antisense oligonucleotides can regulate gene expression in living cells. As such, they regulate cell function and division, and can modulate cellular responses to internal and external stresses and stimuli. Although encouraging results from preclinical and clinical studies have been obtained and significant progress has been made in developing these agents as drugs, they are not yet recognized as effective therapeutics. Several major hurdles remain to be overcome, including problems with efficacy, off-target effects, delivery and side effects. The lessons learned from antisense drug development can help in the development of other oligonucleotide-based therapeutics such as CpG oligonucleotides, RNAi and miRNA. - doi:10.1016/j.drudis.2008.03.014 - 2008

PubReading [16] - Measuring cancer evolution from the genome - T. Graham and A. Sottoriva -

Mon, 30 Aug 2021 18:51:17 GMT

The temporal dynamics of cancer evolution remain elusive, because it is impractical to longitudinally observe cancers unperturbed by treatment. Consequently, our knowledge of how cancers grow largely derives from inferences made from a single point in time – the endpoint in the cancer’s evolution, when it is removed from the body and studied in the laboratory. Fortuitously however, the cancer genome, by virtue of ongoing mutations that uniquely mark clonal lineages within the tumour, provides a rich, yet surreptitious, record of cancer development. In this review, we describe how a cancer’s genome can be analysed to reveal the temporal history of mutation and selection, and discuss why both selective and neutral evolution feature prominently in carcinogenesis. We argue that selection in cancer can only be properly studied once we have some understanding of what the absence of selection looks like. We review the data describing punctuated evolution in cancer, and reason that punctuated phenotype evolution is consistent with both gradual and punctuated genome evolution. We conclude that, to map and predict evolutionary trajectories during carcinogenesis, it is critical to better understand the relationship between genotype change and phenotype change.- DOI: 10.1002/path.4821 - 2016

PubReading [15] - Engineering antibody therapeutics - M. Chiu and G. Gilliland

Tue, 24 Aug 2021 00:30:34 GMT

The successful introduction of antibody-based protein therapeutics into the arsenal of treatments for patients has within a few decades fostered intense innovation in the production and engineering of antibodies. Reviewed here are the methods currently used to produce antibodies along with how our knowledge of the structural and functional characterization of immunoglobulins has resulted in the engineering of antibodies to produce protein therapeutics with unique properties, both biological and biophysical, that are leading to novel therapeutic approaches. Antibody engineering includes the introduction of the antibody combining site (variable regions) into a host of architectures including bi and multi-specific formats that further impact the therapeutic properties leading to further advantages and successes in patient treatment. - http://dx.doi.org/10.1016/j.sbi.2016.07.012 - 2016

PubReading [13] - Herpesvirus capsid assembly and DNA packaging - J. Heming, J. Conway and F. Homa

Thu, 19 Aug 2021 23:30:47 GMT

Herpes simplex virus type I (HSV-1) is the causative agent of several pathologies ranging in severity from the common cold sore to life-threatening encephalitic infection. During productive lytic infection, over 80 viral proteins are expressed in a highly regulated manner, resulting in the replication of viral genomes and assembly of progeny virions. The virion of all herpesviruses consists of an external membrane envelope, a proteinaceous layer called the tegument, and an icosahedral capsid containing the double-stranded linear DNA genome. The capsid shell of HSV-1 is built from four structural proteins: a major capsid protein, VP5, which forms the capsomers (hexons and pentons), the triplex consisting of VP19C and VP23 found between the capsomers, and VP26 which binds to VP5 on hexons but not pentons. In addition, the dodecameric pUL6 portal complex occupies one of the 12 capsid vertices, and the capsid vertex specific component (CVSC), a heterotrimer complex of pUL17, pUL25 and pUL36 binds specifically to the triplexes adjacent to each penton. The capsid is assembled in the nucleus where the viral genome is packaged into newly assembled closed capsid shells. Cleavage and packaging of replicated, concatemeric viral DNA requires the seven viral proteins encoded by the UL6, UL15, UL17, UL25, UL28, UL32, and UL33 genes. Considerable advances have been made in understanding the structure of the herpesvirus capsid and the function of several of the DNA packaging proteins by applying biochemical, genetic, and structural techniques. This review is a summary of recent advances with respect to the structure of the HSV-1 virion capsid and what is known about the function of the seven packaging proteins and their interactions with each other and with the capsid shell. - doi:10.1007/978-3-319-53168-7_6. - 2017

PubReading [12] - Ruthenium Complexes as Anticancer Agents: A Brief History and Perspectives - S. Lee, C. Kim, T.G. Nam

Wed, 18 Aug 2021 21:53:14 GMT

Platinum (Pt)-based anticancer drugs such as cisplatin have been used to treat various cancers. However, they have some limitations including poor selectivity and toxicity towards normal cells and increasing chemoresistance. Therefore, there is a need for novel metallo-anticancers, which has not been met for decades. Since the initial introduction of ruthenium (Ru) polypyridyl complex, a number of attempts at structural evolution have been conducted to improve efficacy. Among them, half-sandwich Ru-arene complexes have been the most prominent as an anticancer platform. Such complexes have clearly shown superior anticancer profiles such as increased selectivity toward cancer cells and ameliorating toxicity against normal cells compared to existing Pt-based anticancers. Currently, several Ru complexes are under human clinical trials. For improvement in selectivity and toxicity associated with chemotherapy, Ru complexes as photodynamic therapy (PDT), and photoactivated chemotherapy (PACT), which can selectively activate prodrug moieties in a specific region, have also been investigated. With all these studies on these interesting entities, new metallo- anticancer drugs to at least partially replace existing Pt-based anticancers are anticipated. This review covers a brief description of Ru-based anticancer complexes and perspectives. https://doi.org/10.2147/DDDT.S275007 - 2020

PubReading [9] - Principles of ubiquitin and SUMO modifications in DNA repair - S. Bergink, S. Jentsch

Fri, 13 Aug 2021 23:31:00 GMT

With the discovery in the late 1980s that the DNA-repair gene RAD6 encodes a ubiquitin-conjugating enzyme, it became clear that protein modification by ubiquitin conjugation has a much broader significance than had previously been assumed. Now, two decades later, ubiquitin and its cousin SUMO are implicated in a range of human diseases, including breast cancer and Fanconi anaemia, giving fresh momentum to studies focused on the relationships between ubiquitin, SUMO and DNA-repair pathways. - NATURE - 2009 -. doi:10.1038/nature07963

PubReading [8] - A Glimpse of Structural Biology through X-Ray Crystallography - Yigong Shi

Fri, 06 Aug 2021 17:55:53 GMT

Since determination of the myoglobin structure in 1957, X-ray crystallography, as the anchoring tool of structural biology, has played an instrumental role in deciphering the secrets of life. Knowledge gained through X-ray crystallography has fundamentally advanced our views on cellular processes and greatly facilitated development of modern medicine. In this brief narrative, I describe my personal understanding of the evolution of structural biology through X-ray crystallography—using as examples mechanistic understanding of protein kinases and integral membrane proteins—and comment on the impact of technological development and outlook of X-ray crystallography - dx.doi.org/10.1016/j.cell.2014.10.051 - 2014

PubReading [7] - Modified nucleic acids: replication, evolution, and next-generation therapeutics - K. Duffy, S. Arangundy-Franklin and P. Holliger

Thu, 05 Aug 2021 23:04:16 GMT

Modified nucleic acids, also called xeno nucleic acids (XNAs), offer a variety of advantages for biotechnological applications and address some of the limitations of first-generation nucleic acid therapeutics. Indeed, several therapeutics based on modified nucleic acids have recently been approved and many more are under clinical evaluation. XNAs can provide increased biostability and furthermore are now increasingly amenable to in vitro evolution, accelerating lead discovery. Here, we review the most recent discoveries in this dynamic field with a focus on progress in the enzymatic replication and functional exploration of XNAs. K. Duffy, S. Arangundy-Franklin and P. Holliger -doi.org/10.1186/s12915-020-00803-6 - 2020

PubReading [250] - Promoting Scientist–Advocate Collaborations in Cancer Research- Why and How -

Mon, 26 Dec 2022 12:00:25 GMT

Advocates bring unique and important viewpoints to the cancer research process, ensuring that scientific and medical advances are patient-centered and relevant. In this article, we discuss the benefits of engaging advocates in cancer research and underscore ways in which both the scientific and patient communities can facilitate this mutually beneficial collaboration. We discuss how to establish and nurture successful scientist–advocate relationships throughout the research process. We review opportunities that are available to advocates who want to obtain training in the evaluation of cancer research. We also suggest practical solutions that can strengthen communication between scientists and advocates, such as introducing scientist–advocate interactions at the trainee level. Finally, we highlight the essential role social media can play in disseminating patient-supported cancer research findings to the patient community and in raising awareness of the importance of promoting cancer research. Our perspective offers a model that Georgetown Breast Cancer Advocates have found effective and which could be one option for those interested in developing productive, successful, and sustainable collaborations between advocates and scientists in cancer research.

doi:10.1158/0008-5472.CAN-18-1600 - 2018

PubReading [6] Will CRISPR‐Cas9 Have Cards to Play Against Cancer? An Update on its Applications - P Daisy, K Shreyas, T. S. Anitha - Molecular Biotechnology

Tue, 03 Aug 2021 00:08:16 GMT

Genome editing employs targeted nucleases as powerful tools to precisely alter the genome of target cells and regulate functional genes. Various strategies have been risen so far as the molecular scissors-mediated genome editing that includes zinc finger nuclease, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats—CRISPR-related protein 9. These tools allow researchers to understand the basics of manipulating the genome, create animal models to study human diseases, understand host–pathogen interactions and design disease targets. Targeted genome modification utilizing RNA-guided nucleases are of recent curiosity, as it is a fast and effective strategy that enables the researchers to manipulate the gene of interest, carry out functional studies, understand the molecular basis of the disease and design targeted therapies. CRISPR-Cas9, a bacterial defense system employed against viruses, consists of a single-strand RNA-guided Cas9 nuclease connected to the corresponding complementary target sequence. This powerful and versatile tool has gained tremendous attention among the researchers, owing to its ability to correct genetic disorders. To help illus- trate the potential of this gene editor in unexplored corners of oncology, we describe the history of CRISPR-Cas9, its rapid progression in cancer research as well as future perspectives. - P Daisy, K Shreyas, T. S. Anitha -doi.org/10.1007/s12033-020-00289-1 - 2021

PubReading [245] - The biospheric emergency calls for scientists to change tactics - F. Racimo, J. Halder et al.

Mon, 19 Dec 2022 12:00:12 GMT

Our current economic and political structures have an increasingly devastating impact on the Earth’s climate and ecosystems: we are facing a biospheric emergency, with catastrophic consequences for both humans and the natural world on which we depend. Life scientists – including biologists, medical scientists, psychologists and public health experts – have had a crucial role in documenting the impacts of this emergency, but they have failed to drive governments to take action in order to prevent the situation from getting worse. Here we, as members of the movement Scientist Rebellion, call on life scientists to re-embrace advocacy and activism – which were once hallmarks of academia – in order to highlight the urgency and necessity of systemic change across our societies. We particularly emphasise the need for scientists to engage in nonviolent civil resistance, a form of public engagement which has proven to be highly effective in social struggles throughout history.

DOI: https://doi.org/10.7554/eLife.83292 - 2022

PubReading [5] G-Quadruplexes at Telomeres: Friend or Foe? - Tracy Bryan - Molecules

Wed, 28 Jul 2021 22:05:50 GMT

Telomeres are DNA-protein complexes that cap and protect the ends of linear chromosomes. In almost all species, telomeric DNA has a G/C strand bias, and the short tandem repeats of the G-rich strand have the capacity to form into secondary structures in vitro, such as four-stranded G-quadruplexes. This has long prompted speculation that G-quadruplexes play a positive role in telomere biology, resulting in selection for G-rich tandem telomere repeats during evolution. There is some evidence that G-quadruplexes at telomeres may play a protective capping role, at least in yeast, and that they may positively affect telomere maintenance by either the enzyme telomerase or by recombination-based mechanisms. On the other hand, G-quadruplex formation in telomeric DNA, as elsewhere in the genome, can form an impediment to DNA replication and a source of genome instability. This review summarizes recent evidence for the in vivo existence of G-quadruplexes at telomeres, with a focus on human telomeres, and highlights some of the many unanswered questions regarding the location, form, and functions of these structures. - Tracy Bryan - doi:10.3390/molecules25163686 - 2020

PubReading [4] Triplex-forming oligonucleotides: a third strand for DNA nanotechnology - Arun Chandrasekaran, David Rusling - Nucleic Acids Research

Tue, 27 Jul 2021 00:26:53 GMT

DNA self-assembly has proved to be a useful bottom- up strategy for the construction of user-defined nanoscale objects, lattices and devices. The design of these structures has largely relied on exploiting simple base-pairing rules and the formation of double-helical domains as secondary structural elements. However, other helical forms involving specific non-canonical base-base interactions have introduced a novel paradigm into the process of engineering with DNA. The most notable of these is a three-stranded complex generated by the binding of a third strand within the duplex major groove, generating a triple-helical (‘triplex’) structure. The sequence, structural and assembly requirements that differentiate triplexes from their duplex counterparts has allowed the design of nanostructures for both dynamic and/or structural purposes, as well as a means to target non-nucleic acid components to precise locations within a nanostructure scaffold. Here, we review the properties of triplexes that have proved useful in the engineering of DNA nanostructures, with an emphasis on applications that hitherto have not been possible by duplex formation alone. Arun Chandrasekaran, David Rusling - doi: 10.1093/nar/gkx1230 - 2018

PubReading [3] DNA Structure: A-, B- and Z-DNA Helix Families - David W Ussery - Encyclopedia Of Life Science

Thu, 22 Jul 2021 23:30:22 GMT

There are three major families of DNA helices: A-DNA, B-DNA and Z-DNA. The helical

structure of DNA is variable and depends on the sequence as well as the environment. David W Ussery

PubReading [2] New Approaches Towards Recognition of Nucleic Acid Triple Helices - Dev P. Arya - Acc Chem Res

Tue, 20 Jul 2021 23:32:12 GMT

We show that groove recognition of nucleic acid triple helices can be achieved with aminosugars. Among these aminosugars, neomycin is the most effective aminoglycoside (groove binder) for stabilizing a DNA triple helix. It stabilizes both the T·A·T triplex and mixed-base DNA triplexes better than known DNA minor groove binders (which usually destabilize the triplex) and polyamines. Neomycin selectively stabilizes the triplex (T·A·T and mixed base) without any effect on the DNA duplex. The selectivity of neomycin likely originates from its potential and shape complementarity to the triplex Watson–Hoogsteen groove, making it the first molecule that selectively recognizes a triplex groove over a duplex groove. The groove recognition of aminoglycosides is not limited to DNA triplexes, but also extends to RNA and hybrid triple helical structures. Intercalator–neomycin conjugates are shown to simultaneously probe the base stacking and groove surface in the DNA triplex. Calorimetric and spectrosocopic studies allow the quantification of the effect of surface area of the intercalating moiety on binding to the triplex. These studies outline a novel approach to the recognition of DNA triplexes that incorporates the use of non-competing binding sites. These principles of dual recognition should be applicable to the design of ligands that can bind any given nucleic acid target with nanomolar affinities and with high selectivity. Dev P. Arya

PubReading [1] A structure for Deoxyribose Nucleic Acid - J. D. Watson, F. H. C. Crick - Nature

Sat, 17 Jul 2021 12:16:38 GMT

Molecular Structure of Nucleic Acids - A structure for Deoxyribose Nucleic Acid - We wish to suggest astructure for the salt of deoxyribose nucleic acid (D.N.A.). This structure has novel features which are of considerable biological interest. J. D. Watson, F. H. C Crick.