Rxivist combines preprints from bioRxiv with data from Twitter to help you find the papers being discussed in your field. Currently indexing 92,757 bioRxiv papers from 396,049 authors.
Most downloaded bioRxiv papers, since beginning of last month
90,515 results found. For more information, click each entry to expand.
475 downloads animal behavior and cognition
Although extensive behavioral changes often exist between closely related animal species, our understanding of the genetic basis underlying the evolution of behavior has remained limited. Here, we propose a new framework to study behavioral evolution by computational estimation of ancestral behavioral repertoires. We measured the behaviors of individuals from six species of fruit flies using unsupervised techniques and identified suites of stereotyped movements exhibited by each species. We then fit a Generalized Linear Mixed Model to estimate the suites of behaviors exhibited by ancestral species, as well as the intra- and inter-species behavioral covariances. We found that much of intraspecific behavioral variation is explained by differences between individuals in the status of their behavioral hidden states, what might be called their "mood." Lastly, we propose a method to identify groups of behaviors that appear to have evolved together, illustrating how sets of behaviors, rather than individual behaviors, likely evolved. Our approach provides a new framework for identifying co-evolving behaviors and may provide new opportunities to study the genetic basis of behavioral evolution. ### Competing Interest Statement The authors have declared no competing interest.
475 downloads developmental biology
During development, embryonic tissues are formed by the dynamic behaviours of their constituent cells, whose collective actions are tightly regulated in space and time. To understand such cell behaviours and how they have evolved, it is necessary to develop quantitative approaches to map out morphogenesis, so comparisons can be made across different tissues and organisms. With this idea in mind, here we sought to investigate ancestral principles of notochord development, by building a quantitative portrait of notochord morphogenesis in the amphioxus embryo, a basally-branching member of the chordate phylum. To this end, we developed a single-cell morphometrics pipeline to comprehensively catalogue the morphologies of thousands of notochord cells, and to project them simultaneously into a common mathematical space termed morphospace. This approach revealed complex patterns of cell-type specific shape trajectories, akin to those obtained using single-cell genomic approaches. By spatially mapping single-cell shape trajectories in whole segmented notochords, we found evidence of spatial and temporal variation in developmental dynamics. Such variations included temporal gradients of morphogenesis across the anterior-posterior embryonic axis, divergence of trajectories to different morphologies, and the convergence of different trajectories onto common morphologies. Through geometric simulations, we also identified an antagonistic relationship between cell shape regulation and growth that enables convergent extension to occur in two steps. First, by allowing growth to counterbalance loss of anterior-posterior cell length during cell intercalation. Secondly, by allowing growth to further increase cell length once cells have intercalated and aligned to the axial midline, thereby facilitating a second phase of tissue elongation. Finally, we show that apart from a complex coordination of individual cellular behaviours, posterior addition from proliferating progenitors is essential for full notochord elongation in amphioxus, a mechanism previously described only in vertebrates. This novel approach to quantifying morphogenesis paves the way towards comparative studies, and mechanistic explanations for the emergence of form over developmental and evolutionary time scales. ### Competing Interest Statement The authors have declared no competing interest.
473 downloads neuroscience
The Assay for Transposase Accessible Chromatin by sequencing (ATAC-seq) is becoming increasingly popular in the neuroscience field where chromatin regulation is thought to be involved in neurodevelopment, activity-dependent gene regulation, hormonal and environmental responses, and the pathophysiology of neuropsychiatric disorders. The advantages of using this assay include a small amount of material needed, relatively simple and fast protocol, and the ability to capture a range of gene regulatory elements with a single assay. However, with increasing interest in chromatin research, it is an imperative to have feasible, reliable assays that are compatible with a range of neuroscience study designs in both animals and humans. Here we tested three different protocols for neuronal chromatin accessibility analysis, including a varying brain tissue freezing method followed by fluorescent-activated nuclei sorting (FANS) and the ATAC-seq analysis. Our study shows that the cryopreservation method impacts the number of open chromatin regions that can be identified from frozen brain tissue using the cell-type specific ATAC-seq assay. However, we show that all three protocols generate consistent and robust data and enable the identification of functional regulatory elements, promoters and enhancers, in neuronal cells. Our study also implies that the broad biological interpretation of chromatin accessibility data is not significantly affected by the freezing condition. In comparison to the mouse brain analysis, we reveal the additional challenges of doing chromatin analysis on post mortem human brain tissue. However, we also show that these studies are revealing important cell type-specific information about gene regulation in the human brain. Overall, the ATAC-seq coupled with FANS is a powerful method to capture cell-type specific chromatin accessibility information in the mouse and human brain. Our study provides alternative brain preservation methods that generate high quality ATAC-seq data while fitting in different study designs, and further encourages the use of this method to uncover the role of epigenetic (dys)regulation in healthy and malfunctioning brain. ### Competing Interest Statement The authors have declared no competing interest.
473 downloads plant biology
Reference datasets are critical in computational biology. They help define canonical biological features and are essential for benchmarking studies. Here, we describe a comprehensive reference dataset of experimentally validated plant NLR immune receptors. RefPlantNLR consists of 415 NLRs from 31 genera belonging to 11 orders of flowering plants. We used RefPlantNLR to determine the canonical features of functionally validated plant NLRs. This reference dataset should prove useful for benchmarking NLR annotation tools, guiding comparative analyses of NLRs across the wide spectrum of plant diversity and identifying under-studied taxa. We hope that the RefPlantNLR resource will contribute to moving the field beyond a uniform view of NLR structure and function. ### Competing Interest Statement The authors receive funding from industry on NLR biology.
473 downloads systems biology
Katie Heiser, Peter F McLean, Chadwick T Davis, Ben Fogelson, Hannah B. Gordon, Pamela Jacobson, Brett Hurst, Ben Miller, Ronald W. Alfa, Berton A. Earnshaw, Mason L. Victors, Yolanda T. Chong, Imran S Haque, Adeline S. Low, Christopher C Gibson
To identify potential therapeutic stop-gaps for SARS-CoV-2, we evaluated a library of 1,670 approved and reference compounds in an unbiased, cellular image-based screen for their ability to suppress the broad impacts of the SARS-CoV-2 virus on phenomic profiles of human renal cortical epithelial cells using deep learning. In our assay remdesivir is the only antiviral tested with strong efficacy, that neither chloroquine nor hydroxychloroquine have any beneficial effect in this human cell model, and that a small number of compounds not currently being pursued clinically for SARS-CoV-2 have efficacy. We observed weak but beneficial class effects of 𝛃-blockers, mTOR/PI3K inhibitors and Vitamin D analogues and a mild amplification of the viral phenotype with 𝛃-agonists. ### Competing Interest Statement All authors from Recursion have real or potential ownership interest in the company. However, Recursion has committed to free non-discriminatory licensing for any of its intellectual property around discoveries related to the treatment of COVID19.
472 downloads microbiology
The outbreak of a novel betacoronavirus (2019-nCov) represents a pandemic threat that has been declared a public health emergency of international concern. The CoV spike (S) glycoprotein is a key target for urgently needed vaccines, therapeutic antibodies, and diagnostics. To facilitate medical countermeasure (MCM) development we determined a 3.5 Å-resolution cryo-EM structure of the 2019-nCoV S trimer in the prefusion conformation. The predominant state of the trimer has one of the three receptor-binding domains (RBDs) rotated up in a receptor-accessible conformation. We also show biophysical and structural evidence that the 2019-nCoV S binds ACE2 with higher affinity than SARS-CoV S. Additionally we tested several published SARS-CoV RBD-specific monoclonal antibodies and found that they do not have appreciable binding to nCoV-2019 S, suggesting antibody cross-reactivity may be limited between the two virus RBDs. The cryo-EM structure of 2019-nCoV S should enable rapid development and evaluation of MCMs to address the ongoing public health crisis.
472 downloads microbiology
Stephen R. Welch, Katherine A. Davies, Hubert Buczkowski, Nipunadi Hettiarachchi, Nicole Green, Ulrike Arnold, Matthew Jones, Matthew J Hannah, Reah Evans, Christopher Burton, Jane E Burton, Malcolm Guiver, Patricia A Cane, Neil Woodford, Christine B Bruce, Allen D. G. Roberts, Marian J. Killip
The COVID-19 pandemic has necessitated a multi-faceted rapid response by the scientific community, bringing researchers, health officials and industry together to address the ongoing public health emergency. To meet this challenge, participants need an informed approach for working safely with the etiological agent, the novel human coronavirus SARS-CoV-2. Work with infectious SARS-CoV-2 is currently restricted to high-containment laboratories, but material can be handled at a lower containment level after inactivation. Given the wide array of inactivation reagents that are being used in laboratories during this pandemic, it is vital that their effectiveness is thoroughly investigated. Here, we evaluated a total of 23 commercial reagents designed for clinical sample transportation, nucleic acid extraction and virus inactivation for their ability to inactivate SARS-CoV-2, as well as seven other common chemicals including detergents and fixatives. As part of this study, we have also tested five filtration matrices for their effectiveness at removing the cytotoxic elements of each reagent, permitting accurate determination of levels of infectious virus remaining following treatment. In addition to providing critical data informing inactivation methods and risk assessments for diagnostic and research laboratories working with SARS-CoV-2, these data provide a framework for other laboratories to validate their inactivation processes and to guide similar studies for other pathogens.
472 downloads microbiology
Chee Keng Mok, Yan Ling Ng, Bintou Ahmadou Ahidjo, Regina Ching Hua Lee, Marcus Wing Choy Loe, Jing Liu, Kai Sen Tan, Parveen Kaur, Wee Joo Chng, John Eu-Li Wong, De Yun Wang, Erwei Hao, Xiaotao Hou, Yong Wah Tan, Tze Minn Mak, Cui Lin, Raymond Lin, Paul Tambyah, JiaGang Deng, Justin Jang Hann Chu
COVID-19, the disease caused by SARS-CoV-2, was declared a pandemic by the World Health Organization (WHO) in March 2020. While awaiting a vaccine, several antivirals are being used to manage the disease with limited success. To expand this arsenal, we screened 4 compound libraries: a United States Food and Drug Administration (FDA) approved drug library, an angiotensin converting enzyme-2 (ACE2) targeted compound library, a flavonoid compound library as well as a natural product library. Of the 121 compounds identified with activity against SARS-CoV-2, 7 were shortlisted for validation. We show for the first time that the active form of Vitamin D, calcitriol, exhibits significant potent activity against SARS-CoV-2. This finding paves the way for consideration of host-directed therapies for ring prophylaxis of contacts of SARS-CoV-2 patients. ### Competing Interest Statement The authors have declared no competing interest.
472 downloads immunology
Seth J Zost, Pavlo Gilchuk, James Brett Case, Elad Binshtein, Rita E. Chen, Joseph X Reidy, Andrew Trivette, Rachel S Nargi, Rachel E Sutton, Naveenchandra Suryadevara, Lauren E Williamson, Elaine C Chen, Taylor Jones, Samuel Day, Luke Myers, Ahmed O Hassan, Natasha M. Kafai, Emma S Winkler, Julie M Fox, James J Steinhardt, Kuishu Ren, Yueh-Ming Loo, Nicole L Kallewaard, David R. Martinez, Alexandra Schäfer, Lisa E. Gralinski, Ralph S. Baric, Larissa B Thackray, Michael S. Diamond, Robert H. Carnahan, James E. Crowe
The COVID-19 pandemic is a major threat to global health for which there are only limited medical countermeasures, and we lack a thorough understanding of mechanisms of humoral immunity. From a panel of monoclonal antibodies (mAbs) targeting the spike (S) glycoprotein isolated from the B cells of infected subjects, we identified several mAbs that exhibited potent neutralizing activity with IC50 values as low as 0.9 or 15 ng/mL in pseudovirus or wild-type (wt) SARS-CoV-2 neutralization tests, respectively. The most potent mAbs fully block the receptor-binding domain of S (SRBD) from interacting with human ACE2. Competition-binding, structural, and functional studies allowed clustering of the mAbs into defined classes recognizing distinct epitopes within major antigenic sites on the SRBD. Electron microscopy studies revealed that these mAbs recognize distinct conformational states of trimeric S protein. Potent neutralizing mAbs recognizing unique sites, COV2-2196 and COV2-2130, bound simultaneously to S and synergistically neutralized authentic SARS-CoV-2 virus. In two murine models of SARS-CoV-2 infection, passive transfer of either COV2-2916 or COV2-2130 alone or a combination of both mAbs protected mice from severe weight loss and reduced viral burden and inflammation in the lung. These results identify protective epitopes on the SRBD and provide a structure-based framework for rational vaccine design and the selection of robust immunotherapeutic cocktails. ### Competing Interest Statement R.S.B. has served as a consultant for Takeda and Sanofi Pasteur on issues related to vaccines. M.S.D. is a consultant for Inbios, Vir Biotechnology, NGM Biopharmaceuticals, Eli Lilly, and is on the Scientific Advisory Board of Moderna, a past recipient of unrelated research grant from Moderna and a current recipient of an unrelated research grant Emergent BioSolutions. J.E.C. has served as a consultant for Sanofi and is on the Scientific Advisory Boards of CompuVax and Meissa Vaccines, is a recipient of previous unrelated research grants from Moderna and Sanofi and is Founder of IDBiologics, Inc. Vanderbilt University has applied for patents concerning SARS-CoV-2 antibodies that are related to this work. AstraZeneca has filed patents for materials/findings related to this work. J.J.S., K.R., Y.-M.L., and N.L.K. are employees of AstraZeneca and currently hold AstraZeneca stock or stock options. All other authors declared no competing interests.
469 downloads immunology
Shuo Du, Yunlong Cao, Qinyu Zhu, Guopeng Wang, Xiaoxia Du, Runsheng He, Hua Xu, Yinghui Zheng, Bo Wang, Yali Bai, Chenggong Ji, Ayijiang Yisimayi, Qisheng Wang, Ning Gao, X. Sunney Xie, Xiao-dong Su, Junyu Xiao
Understanding the mechanism of neutralizing antibodies (NAbs) against SARS-CoV-2 is critical for effective vaccines and therapeutics development. We recently reported an exceptionally potent NAb, BD-368-2, and revealed the existence of VH3-53/VH3-66 convergent NAbs in COVID-19. Here we report the 3.5 angstrom cryo-EM structure of the Fabs of BD-368-2 in complex with a mutation-induced prefusion-state-stabilized spike trimer. Unlike VH3-53/VH3-66 NAbs, BD-368-2 fully blocks ACE2 binding by occupying all three receptor-binding domains (RBDs) simultaneously, regardless of their up and down positions. BD-368-2 also triggers fusogenic-like structural rearrangements of the spike trimer, which could impede viral entry. Moreover, BD-368-2 completely avoids the common epitope of VH3-53/VH3-66 NAbs, evidenced by multiple crystal structures of their Fabs in tripartite complexes with RBD, suggesting a new way of pairing potent NAbs to prevent neutralization escape. Together, these results rationalize a unique epitope that leads to exceptional neutralization potency, and provide guidance for NAb therapeutics and vaccine designs against SARS-CoV-2. ### Competing Interest Statement X.S.X and Y.C are inventors on the patent applications of the NAbs.
469 downloads molecular biology
Matthias Thoms, Robert Buschauer, Michael Ameismeier, Lennart Koepke, Timo Denk, Maximilian Hirschenberger, Hanna Kratzat, Manuel Hayn, Timur Mackens-Kiani, Jingdong Cheng, Christina M. Stürzel, Thomas Fröhlich, Otto Berninghausen, Thomas Becker, Frank Kirchhoff, Konstantin M.J. Sparrer, Roland Beckmann
SARS-CoV-2 is the causative agent of the current COVID-19 pandemic. A major virulence factor of SARS-CoVs is the nonstructural protein 1 (Nsp1) which suppresses host gene expression by ribosome association via an unknown mechanism. Here, we show that Nsp1 from SARS-CoV-2 binds to 40S and 80S ribosomes, resulting in shutdown of capped mRNA translation both in vitro and in cells. Structural analysis by cryo-electron microscopy (cryo-EM) of in vitro reconstituted Nsp1-40S and of native human Nsp1-ribosome complexes revealed that the Nsp1 C-terminus binds to and obstructs the mRNA entry tunnel. Thereby, Nsp1 effectively blocks RIG-Idependent innate immune responses that would otherwise facilitate clearance of the infection. Thus, the structural characterization of the inhibitory mechanism of Nsp1 may aid structure-based drug design against SARS-CoV-2. ### Competing Interest Statement The authors have declared no competing interest.
469 downloads developmental biology
The evolutionary mechanisms underlying the emergence of new cell types are still unclear. Here, we address the origin and diversification of muscle cells in the diploblastic sea anemone Nematostella vectensis. We discern two fast and two slow-contracting muscle cell populations in Nematostella differing by extensive sets of paralogous genes. The regulatory gene set of the slow cnidarian muscles and the bilaterian cardiac muscle are remarkably similar. By contrast, the two fast muscles differ substantially from each other, while driving the same set of paralogous structural protein genes. Our data suggest that extensive gene duplications and co-option of individual effector modules may have played an important role in cell type diversification during metazoan evolution. ### Competing Interest Statement The authors have declared no competing interest.
468 downloads immunology
Reuben McGregor, Daniel Chauss, Tilo Freiwald, Bingyu Yan, Luopin Wang, Estefania Nova-Lamperti, Zonghao Zhang, Heather Teague, Erin E West, Jack Bibby, Audrey Kelly, Amna Malik, Alexandra F. Freeman, Daniella Schwartz, Didier Portilla, Susan John, Paul Lavender, Michail S Lionakis, Nehal N Mehta, Claudia Kemper, Nichola Cooper, Giovanna Lombardi, Arian Laurence, M. Kazemian, Behdad Afzali
Pro-inflammatory immune responses are necessary for effective pathogen clearance, but cause severe tissue damage if not shut down in a timely manner. Excessive complement and IFN-γ-associated responses are known drivers of immunopathogenesis and are among the most highly induced immune programs in hyper-inflammatory SARS-CoV2 lung infection. The molecular mechanisms that govern orderly shutdown and retraction of these responses remain poorly understood. Here, we show that complement triggers contraction of IFN-γ producing CD4+ T helper (Th) 1 cell responses by inducing expression of the vitamin D (VitD) receptor (VDR) and CYP27B1, the enzyme that activates VitD, permitting T cells to both activate and respond to VitD. VitD then initiates the transition from pro-inflammatory IFN-γ+ Th1 cells to suppressive IL-10+ Th1 cells. This process is primed by dynamic changes in the epigenetic landscape of CD4+ T cells, generating super-enhancers and recruiting c-JUN and BACH2, a key immunoregulatory transcription factor. Accordingly, cells in psoriatic skin treated with VitD increased BACH2 expression, and BACH2 haplo-insufficient CD4+ T cells were defective in IL-10 production. As proof-of-concept, we show that CD4+ T cells in the bronchoalveolar lavage fluid (BALF) of patients with COVID-19 are Th1-skewed and that VDR is among the top regulators of genes induced by SARS-CoV2. Importantly, genes normally down-regulated by VitD were de-repressed in CD4+ BALF T cells of COVID-19, indicating that the VitD-driven shutdown program is impaired in this setting. The active metabolite of VitD, alfacalcidol, and cortico-steroids were among the top predicted pharmaceuticals that could normalize SARS-CoV2 induced genes. These data indicate that adjunct therapy with VitD in the context of other immunomodulatory drugs may be a beneficial strategy to dampen hyper-inflammation in severe COVID-19. ### Competing Interest Statement The authors have declared no competing interest.
468 downloads neuroscience
Decades of work have demonstrated that mRNAs are localized and translated within neuronal dendrites and axons to provide proteins for remodeling and maintaining growth cones or synapses. It remains unknown, however, whether specific forms of plasticity differentially regulate the dynamics and translation of individual mRNA species. To address these issues, we targeted three individual synaptically-localized mRNAs, CamkIIa, Beta actin, Psd95, and used molecular beacons to track endogenous mRNA movements and reporters and Crispr-Cas9 gene editing to track their translation. We found widespread alterations in mRNA behavior during two forms of synaptic plasticity, long-term potentiation (LTP) and depression (LTD). Changes in mRNA dynamics following plasticity resulted in an enrichment of mRNA in the vicinity of dendritic spines. Both the reporters and tagging of endogenous proteins revealed the transcript-specific stimulation of protein synthesis following LTP or LTD. The plasticity-induced enrichment of mRNA near synapses could be uncoupled from its translational status. The enrichment of mRNA in the proximity of spines allows for localized signaling pathways to decode plasticity milieus and stimulate a specific translational profile, resulting in a customized remodeling of the synaptic proteome. ### Competing Interest Statement The authors have declared no competing interest.
468 downloads immunology
Sonny R. Elizaldi, Yashavanth Shaan Lakshmanappa, Jamin W. Roh, Brian A. Schmidt, Timothy D. Carroll, Kourtney D. Weaver, Justin C. Smith, Jesse D. Deere, Joseph Dutra, Mars Stone, Rebecca Lee Sammak, Katherine J. Olstad, J. Rachel Reader, Zhong-Min Ma, Nancy K. Nguyen, Jennifer Watanabe, Jodie Usachaenko, Ramya Immareddy, JoAnn L. Yee, Daniela Weiskopf, Alessandro Sette, Dennis Hartigan-O’Connor, Stephen J. McSorley, John H Morrison, Nam K Tran, Graham Simmons, Michael P. Busch, Pamela A. Kozlowski, Koen K.A Van Rompay, Christopher J. Miller, Smita S Iyer
CD4 T follicular helper (Tfh) cells are important for the generation of long-lasting and specific humoral protection against viral infections. The degree to which SARS-CoV-2 infection generates Tfh cells and stimulates the germinal center response is an important question as we investigate vaccine options for the current pandemic. Here we report that, following infection with SARS-CoV-2, adult rhesus macaques exhibited transient accumulation of activated, proliferating Tfh cells in their peripheral blood on a transitory basis. The CD4 helper cell responses were skewed predominantly toward a Th1 response in blood, lung, and lymph nodes, reflective of the interferon-rich cytokine environment following infection. We also observed the generation of germinal center Tfh cells specific for the SARS-CoV-2 spike (S) and nucleocapsid (N) proteins, and a corresponding early appearance of antiviral serum IgG antibodies but delayed or absent IgA antibodies. Our data suggest that a vaccine promoting Th1-type Tfh responses that target the S protein may lead to protective immunity. ### Competing Interest Statement A.S. is listed as inventors on a provisional patent application covering findings reported in this manuscript. A.S. is a consultant for Gritstone, Flow Pharma, and Avalia. The other authors have no competing interests to declare.
467 downloads genetics
We create and share a new red fluorophore, along with a set of strains, reagents and protocols, to make it faster and easier to label endogenous C. elegans proteins with fluorescent tags. CRISPR-mediated fluorescent labeling of C. elegans proteins is an invaluable tool, but it is much more difficult to insert fluorophore-size DNA segments than it is to make small gene edits. In principle, high-affinity asymmetrically split fluorescent proteins solve this problem in C. elegans : the small fragment can quickly and easily be fused to almost any protein of interest and can be detected wherever the large fragment is expressed and complemented. There is currently only one available strain stably expressing the large fragment of a split fluorescent protein, restricting this solution to a single tissue (the germline) in the highly autofluorescent green channel. No available C. elegans lines express unbound large fragments of split red fluorescent proteins, and even state-of-the-art split red fluorescent proteins are dim compared to the canonical split-sfGFP protein. In this study, we engineer a bright, high-affinity new split red fluorophore, split-wrmScarlet, and generate transgenic C. elegans lines to allow easy single-color labeling in muscles and dual-color labeling in somatic cells. We validate these strains by targeting split-wrmScarlet to several genes whose products label distinct organelles, and we provide a protocol for an easy, cloning-free method for CRISPR/Cas9 editing. ### Competing Interest Statement The authors have declared no competing interest.
467 downloads systems biology
Emanuel Wyler, Mösbauer Kirstin, Vedran Franke, Diag Asija, Gottula Lina Theresa, Arsie Roberto, Klironomos Filippos, Koppstein David, Ayoub Salah, Buccitelli Christopher, Richter Anja, Legnini Ivano, Ivanov Andranik, Mari Tommaso, Del Giudice Simone, Papies Jan Patrick, Müller Marcel Alexander, Niemeyer Daniela, Selbach Matthias, Altuna Akalin, Nikolaus Rajewsky, Drosten Christian, Markus Landthaler
The coronavirus disease 2019 (COVID-19) pandemic, caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is an ongoing global health threat with more than two million infected people since its emergence in late 2019. Detailed knowledge of the molecular biology of the infection is indispensable for understanding of the viral replication, host responses, and disease progression. We provide gene expression profiles of SARS-CoV and SARS-CoV-2 infections in three human cell lines (H1299, Caco-2 and Calu-3 cells), using bulk and single-cell transcriptomics. Small RNA profiling showed strong expression of the immunity and inflammation-associated microRNA miRNA-155 upon infection with both viruses. SARS-CoV-2 elicited approximately two-fold higher stimulation of the interferon response compared to SARS-CoV in the permissive human epithelial cell line Calu-3, and induction of cytokines such as CXCL10 or IL6. Single cell RNA sequencing data showed that canonical interferon stimulated genes such as IFIT2 or OAS2 were broadly induced, whereas interferon beta (IFNB1) and lambda (IFNL1-4) were expressed only in a subset of infected cells. In addition, temporal resolution of transcriptional responses suggested interferon regulatory factors (IRFs) activities precede that of nuclear factor-κB (NF-κB). Lastly, we identified heat shock protein 90 (HSP90) as a protein relevant for the infection. Inhibition of the HSP90 charperone activity by Tanespimycin/17-N-allylamino-17-demethoxygeldanamycin (17-AAG) resulted in a reduction of viral replication, and of TNF and IL1B mRNA levels. In summary, our study established in vitro cell culture models to study SARS-CoV-2 infection and identified HSP90 protein as potential drug target for therapeutic intervention of SARS-CoV-2 infection. ### Competing Interest Statement The authors have declared no competing interest.
467 downloads microbiology
Neal G. Ravindra, Mia Madel Alfajaro, Victor Gasque, Victoria Habet, Jin Wei, Renata B. Filler, Nicholas C. Huston, Han Wan, Klara Szigeti-Buck, Bao Wang, Guilin Wang, Ruth R. Montgomery, Stephanie C. Eisenbarth, Adam Williams, Anna Marie Pyle, Akiko Iwasaki, Tamas L Horvath, Ellen F. Foxman, Richard W. Pierce, David van Dijk, Craig B Wilen
SARS-CoV-2, the causative agent of COVID-19, has tragically burdened individuals and institutions around the world. There are currently no approved drugs or vaccines for the treatment or prevention of COVID-19. Enhanced understanding of SARS-CoV-2 infection and pathogenesis is critical for the development of therapeutics. To reveal insight into viral replication, cell tropism, and host-viral interactions of SARS-CoV-2 we performed single-cell RNA sequencing of experimentally infected human bronchial epithelial cells (HBECs) in air-liquid interface cultures over a time-course. This revealed novel polyadenylated viral transcripts and highlighted ciliated cells as a major target of infection, which we confirmed by electron microscopy. Over the course of infection, cell tropism of SARS-CoV-2 expands to other epithelial cell types including basal and club cells. Infection induces cell-intrinsic expression of type I and type III IFNs and IL6 but not IL1. This results in expression of interferon-stimulated genes in both infected and bystander cells. We observe similar gene expression changes from a COVID-19 patient ex vivo . In addition, we developed a new computational method termed CONditional DENSity Embedding (CONDENSE) to characterize and compare temporal gene dynamics in response to infection, which revealed genes relating to endothelin, angiogenesis, interferon, and inflammation-causing signaling pathways. In this study, we conducted an in-depth analysis of SARS-CoV-2 infection in HBECs and a COVID-19 patient and revealed genes, cell types, and cell state changes associated with infection. ### Competing Interest Statement The authors have declared no competing interest.
466 downloads microbiology
Fatai S. Oladunni, Jun-Gyu Park, Paula Pino Tamayo, Olga Gonzalez, Anwari Akhter, Anna Allué-Guardia, Angélica Olmo-Fontánez, Shalini Gautam, Andreu Garcia-Vilanova, Chengjin Ye, Kevin Chiem, Colwyn Headley, Varun Dwivedi, Laura M Parodi, Kendra J. Alfson, Hilary M Staples, Alyssa Schami, Juan I. Garcia, Alison Whigham, Roy Neal Platt, Michal Gazi, Jesse Martinez, Colin Chuba, Stephanie Earley, Oscar H Rodriguez, Stephanie Davis Mdaki, Katrina N Kavelish, Renee Escalona, Cory R. A. Hallam, Corbett Christie, Jean L Patterson, Timothy JC. Anderson, Ricardo Carrion, Edward J. Dick, Shannan Hall-Ursone, Larry S. Schlesinger, Deepak Kaushal, Luis D Giavedoni, Xavier Alvarez, Joanne Turner, Luis Martinez-Sobrido, Jordi B. Torrelles
Vaccine and antiviral development against SARS-CoV-2 infection or COVID-19 disease currently lacks a validated small animal model. Here, we show that transgenic mice expressing human angiotensin converting enzyme 2 (hACE2) by the human cytokeratin 18 promoter (K18 hACE2) represent a susceptible rodent model. K18 hACE2-transgenic mice succumbed to SARS-CoV-2 infection by day 6, with virus detected in lung airway epithelium and brain. K18 ACE2-transgenic mice produced a modest TH1/2/17 cytokine storm in the lung and spleen that peaked by day 2, and an extended chemokine storm that was detected in both lungs and brain. This chemokine storm was also detected in the brain at day 4. K18 hACE2-transgenic mice are, therefore, highly susceptible to SARS-CoV-2 infection and represent a suitable animal model for the study of viral pathogenesis, and for identification and characterization of vaccines (prophylactic) and antivirals (therapeutics) for SARS-CoV-2 infection and associated severe COVID-19 disease. ### Competing Interest Statement The authors have declared no competing interest.
465 downloads bioinformatics
Motivation: The alignment of sequencing reads to a transcriptome is a common and important step in many RNA-seq analysis tasks. When aligning RNA-seq reads directly to a transcriptome (as is common in the de novo setting or when a trusted reference annotation is available), care must be taken to report the potentially large number of multi-mapping locations per read. This can pose a substantial computational burden for existing aligners, and can considerably slow downstream analysis. Results: We introduce a novel concept, quasi-mapping, and an efficient algorithm implementing this approach for mapping sequencing reads to a transcriptome. By attempting only to report the potential loci of origin of a sequencing read, and not the base-to-base alignment by which it derives from the reference, RapMap --- our tool implementing quasi-mapping --- is capable of mapping sequencing reads to a target transcriptome substantially faster than existing alignment tools. The algorithm we employ to implement quasi-mapping uses several efficient data structures and takes advantage of the special structure of shared sequence prevalent in transcriptomes to rapidly provide highly-accurate mapping information. We demonstrate how quasi-mapping can be successfully applied to the problems of transcript-level quantification from RNA-seq reads and the clustering of contigs from de novo assembled transcriptomes into biologically-meaningful groups. Availability: RapMap is implemented in C++11 and is available as open-source software, under GPL v3, at https://github.com/COMBINE-lab/RapMap.
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