Biology subjects

Zhu, W.

Publications and source records attributed to Zhu, W..

At least 163 records · Page 9Linked to original sources

bioRxiv2020

Pancreatic α and β cells are globally phase-locked

The Ca2+ modulated pulsatile secretion of glucagon and insulin by pancreatic and {beta} cells plays a key role in glucose homeostasis. However, how and {beta} cells coordinate via paracrine interaction to produce various Ca2+ oscillation patterns is still elusive. Using a microfluidic device and transgenic mice in which and {beta} cells were labeled with different colors, we were able to record islet Ca2+ signals at single cell level for long times. Upon glucose stimulation, we observed heterogeneous Ca2+ oscillation patterns intrinsic to each islet. After a transient period, the oscillations of and {beta} cells were globally phase-locked, i.e., the two types of cells in an islet each oscillate synchronously but with a phase shift between the two. While the activation of cells displayed a fixed time delay of ~20 s to that of {beta} cells, {beta} cells activated with a tunable delay after the cells. As a result, the tunable phase shift between and {beta} cells set the islet oscillation period and pattern. Furthermore, we demonstrated that the phase shift can be modulated by glucagon. A mathematical model of islet Ca2+ oscillation taking into consideration of the paracrine interaction was constructed, which quantitatively agreed with the experimental data. Our study highlights the importance of cell-cell interaction to generate stable but tunable islet oscillation patterns.

systems biology↗

bioRxiv2020

DeepPSC (protein structure camera): computer vision-based reconstruction of proteins backbone structure from alpha carbon trace as a case study

Deep learning has been increasingly used in protein tertiary structure prediction, a major goal in life science. However, all the algorithms developed so far mostly use protein sequences as input, whereas the vast amount of protein tertiary structure information available in the Protein Data Bank (PDB) database remains largely unused, because of the inherent complexity of 3D data computation. In this study, we propose Protein Structure Camera (PSC) as an approach to convert protein structures into images. As a case study, we developed a deep learning method incorporating PSC (DeepPSC) to reconstruct protein backbone structures from alpha carbon traces. DeepPSC outperformed all the methods currently available for this task. This PSC approach provides a useful tool for protein structure representation, and for the application of deep learning in protein structure prediction and protein engineering.

bioinformatics↗

bioRxiv2020

The D614G mutation in the SARS-CoV2 Spike protein increases infectivity in an ACE2 receptor dependent manner

The SARS-CoV2 coronavirus responsible for the current COVID19 pandemic has been reported to have a relatively low mutation rate. Nevertheless, a few prevalent variants have arisen that give the appearance of undergoing positive selection as they are becoming increasingly widespread over time. Most prominent among these is the D614G amino acid substitution in the SARS-CoV2 Spike protein, which mediates viral entry. The D614G substitution, however, is in linkage disequilibrium with the ORF1b P314L mutation where both mutations almost invariably co-occur, making functional inferences problematic. In addition, the possibility of repeated new introductions of the mutant strain does not allow one to distinguish between a founder effect and an intrinsic genetic property of the virus. Here, we synthesized and expressed the WT and D614G variant SARS-Cov2 Spike protein, and report that using a SARS-CoV2 Spike protein pseudotyped lentiviral vector we observe that the D614G variant Spike has >1/2 log10 increased infectivity in human cells expressing the human ACE2 protein as the viral receptor. The increased binding/fusion activity of the D614G Spike protein was corroborated in a cell fusion assay using Spike and ACE2 proteins expressed in different cells. These results are consistent with the possibility that the Spike D614G mutant increases the infectivity of SARS-CoV2.

microbiology↗

bioRxiv2020

Identification of SARS-CoV-2 3CL Protease Inhibitors by a Quantitative High-throughput Screening

The outbreak of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has emphasized the urgency to develop effective therapeutics. Drug repurposing screening is regarded as one of the most practical and rapid approaches for the discovery of such therapeutics. The 3C like protease (3CLpro), or main protease (Mpro) of SARS-CoV-2 is a valid drug target as it is a specific viral enzyme and plays an essential role in viral replication. We performed a quantitative high throughput screening (qHTS) of 10,755 compounds consisting of approved and investigational drugs, and bioactive compounds using a SARS-CoV-2 3CLpro assay. Twenty-three small molecule inhibitors of SARS-CoV-2 3CLpro have been identified with IC50s ranging from 0.26 to 28.85 M. Walrycin B (IC50 = 0.26 {micro}M), Hydroxocobalamin (IC50 = 3.29 {micro}M), Suramin sodium (IC50 = 6.5 {micro}M), Z-DEVD-FMK (IC50 = 6.81 {micro}M), LLL-12 (IC50 = 9.84 {micro}M), and Z-FA-FMK (IC50 = 11.39 {micro}M) are the most potent 3CLpro inhibitors. The activities of anti-SARS-CoV-2 viral infection was confirmed in 7 of 23 compounds using a SARS-CoV-2 cytopathic effect assay. The results demonstrated a set of SARS-CoV-2 3CLpro inhibitors that may have potential for further clinical evaluation as part of drug combination therapies to treating COVID-19 patients, and as starting points for chemistry optimization for new drug development.

pharmacology and toxicology↗

bioRxiv2020

The Botrytis cinerea Crh transglycosylae is a cytoplasmic effector triggering plant cell death and defense response

Crh proteins catalyze crosslinking of chitin and glucan polymers in the fugal cell wall. We revealed a novel and unexpected role of Botrytis cinerea BcCrh1 as a cytoplasmic effector and elicitor of plant defense. During saprophytic growth the BcCrh1 protein is localized in vacuoles and ER. Upon plant infection the protein accumulates to high levels in infection cushions, it is then secreted to the apoplast and translocated into plant cells, where it induces cell death and defense responses. Two regions of 53 and 35 amino acids were found sufficient for protein uptake and cell death induction, respectively. Dimerization of BcCrh proteins was necessary for the transglycosylation activity and proper fungal development, while the monomeric proteins was sufficient for induction of cell death. Arabidopsis lines expressing the bccrh1 gene had reduced sensitivity to B. cinerea, demonstrating the potential use of the protein in plant immunization against necrotrophic pathogens.

microbiology↗

bioRxiv2020

Rapid Inactivation of SARS-CoV-2 by Silicon Nitride, Copper, and Aluminum Nitride

IntroductionViral disease spread by contaminated commonly touched surfaces is a global concern. Silicon nitride, an industrial ceramic that is also used as an implant in spine surgery, has known antibacterial activity. The mechanism of antibacterial action relates to the hydrolytic release of surface disinfectants. It is hypothesized that silicon nitride can also inactivate the coronavirus SARS-CoV-2. MethodsSARS-CoV-2 virions were exposed to 15 wt.% aqueous suspensions of silicon nitride, aluminum nitride, and copper particles. The virus was titrated by the TCD50 method using VeroE6/TMPRSS2 cells, while viral RNA was evaluated by real-time RT-PCR. Immunostaining and Raman spectroscopy were used as additional probes to investigate the cellular responses to virions exposed to the respective materials. ResultsAll three tested materials showed >99% viral inactivation at one and ten minutes of exposure. Degradation of viral RNA was also observed with all materials. Immunofluorescence testing showed that silicon nitride-treated virus failed to infect VeroE6/TMPRSS2 cells without damaging them. In contrast, the copper-treated virus suspension severely damaged the cells due to copper ion toxicity. Raman spectroscopy indicated differential biochemical cellular changes due to infection and metal toxicity for two of the three materials tested. ConclusionsSilicon nitride successfully inactivated the SARS-CoV-2 in this study. The mechanism of action was the hydrolysis-mediated surface release of nitrogen-containing disinfectants. Both aluminum nitride and copper were also effective in the inactivation of the virus. However, while the former compound affected the cells, the latter compound had a cytopathic effect. Further studies are needed to validate these findings and investigate whether silicon nitride can be incorporated into personal protective equipment and commonly touched surfaces, as a strategy to discourage viral persistence and disease spread.

microbiology↗

bioRxiv2020

An OpenData portal to share COVID-19 drug repurposing data in real time

The National Center for Advancing Translational Sciences (NCATS) has developed an online open science platform - named the NCATS OpenData Portal (ODP) - for quickly and freely sharing complete NCATS translational datasets via an open-access, user-friendly interface. This paper describes the establishment of the ODP, initially deployed during the COVID-19 crisis, and provides a detailed analysis of COVID-19 drug repurposing screening datasets that served as the first large-scale use case for the platform. Over 10,000 compounds were tested across 17 quantitative high-throughput assays, covering a wide spectrum of the SARS-CoV-2 life cycle. In total, over 87,000 concentration-response curves and 426,000 data points were made publicly available on ODP in near real-time, enabling immediate access to complete datasets. The resource is flexible in accommodating various types and structures of data, and it has already expanded since its launch to host additional datasets for COVID-19, other viruses of pandemic potential, and beyond. The OpenData Portal has been designed as a scalable platform for real-time data sharing across drug discovery campaigns, regardless of disease area, with the overarching goal of accelerating discovery at NCATS, the NIH, and the greater scientific community.

microbiology↗

bioRxiv2020

The SARS-CoV-2 cytopathic effect is blocked with autophagy modulators

SARS-CoV-2 is a new type of coronavirus capable of rapid transmission and causing severe clinical symptoms; much of which has unknown biological etiology. It has prompted researchers to rapidly mobilize their efforts towards identifying and developing anti-viral therapeutics and vaccines. Discovering and understanding the virus’ pathways of infection, host-protein interactions, and cytopathic effects will greatly aid in the design of new therapeutics to treat COVID-19. While it is known that chloroquine and hydroxychloroquine, extensively explored as clinical agents for COVID-19, have multiple cellular effects including inhibiting autophagy, there are also dose-limiting toxicities in patients that make clearly establishing their potential mechanisms-of-action problematic. Therefore, we evaluated a range of other autophagy modulators to identify an alternative autophagy-based drug repurposing opportunity. In this work, we found that 6 of these compounds blocked the cytopathic effect of SARS-CoV-2 in Vero-E6 cells with EC50 values ranging from 2.0 to 13 µM and selectivity indices ranging from 1.5 to >10-fold. Immunofluorescence staining for LC3B and LysoTracker dye staining assays in several cell lines indicated their potency and efficacy for inhibiting autophagy correlated with the measurements in the SARS-CoV-2 cytopathic effect assay. Our data suggest that autophagy pathways could be targeted to combat SARS-CoV-2 infections and become an important component of drug combination therapies to improve the treatment outcomes for COVID-19.One Sentence Summary Blocking SARS-CoV-2 cytopathic effects with selective autophagy inhibitors underlying the clinical benefits of chloroquine and hydroxychloroquine.Competing Interest StatementThe authors have declared no competing interest.View Full Text

pharmacology and toxicology↗

bioRxiv2020

Alk1 mutant endothelial cells undergo clonal expansion in mouse brain arteriovenous malformations

RationaleMutation in human arteriovenous malformation (AVM) causative genes in a fraction of endothelial cells (ECs) causes AVMs in mice. It is unclear how a small number of mutant ECs can lead to AVM formation. ObjectiveTo understand how a fraction of mutant ECs causes AVM, we tested the following hypotheses: (1) activin receptor-like kinase 1 (Alk1 or Acvlr1) mutant brain ECs undergo clonal expansion upon angiogenic stimulation, (2) Alk1 mutant ECs display growth advantage, (3) the burden of Alk1 mutant ECs correlates with AVM severity, and (4) Alk1 mutant bone marrow (BM) derived ECs alone is sufficient to cause AVM. Methods and ResultsWe used PdgfbiCreER;Alk1f/f;confetti+/- mice which express an EC-specific tamoxifen (TM)-inducible Cre recombinase, a Cre-regulated confetti transgene, and Alk1 floxed alleles. Brain AVMs were induced by direct brain injection of an adeno-associated viral vector expressing vascular endothelial growth factor (AAV-VEGF) followed with intra-peritoneal injection of TM two weeks later. Color-predominance of confetti reporter in AVMs compared to control brain ECs suggested that clonal expansion was associated with AVM development. We treated PdgfbiCreER;Alk1f/f with different doses of TM to create a mosaic of wild-type (WT) and mutant ECs and found that equal numbers of Alk1+ and Alk1- ECs were proliferating. Increase of TM dose increased the number of Alk1- ECs, the abnormal vessels in brain AVMs, the number of arteriovenous shunts in the intestines, and mouse mortality. To test if mutation of Alk1 in BM-derived ECs can cause brain AVM, we transplanted WT mice with BM of PdgfbiCreER;Alk1f/f mice. After AAV-VEGF and TM treatment, these mice developed AVMs in their brains and arteriovenous shunts in their intestines. ConclusionClonal expansion of Alk1 mutant ECs could partly explain why a fraction of mutant ECs causes AVM. Mutation of AVM causal genes in BM-derived ECs is sufficient to cause AVM formation.

cell biology↗

bioRxiv2020

Morphologic Classification and Automatic Nugent Scoring of Bacterial Vaginosis by Deep Neural Networks

BackgroundBacterial vaginosis (BV) was the most common condition for womens health caused by the disruption of normal vaginal flora and an overgrowth of certain disease-causing bacteria, affecting 30-50% of women at some time in their lives. Gram stain followed by Nugent scoring (NS) based on bacterial morphotypes under the microscope was long considered golden standard for BV diagnosis. This conventional manual method was often considered labor intensive, time consuming, and variable results from person to person. MethodsWe developed four convolutional neural networks (CNN) models, and evaluated their ability to automatic identify vaginal bacteria and classify Nugent scores from microscope images. All the CNN models were first trained with 23280 microscopic images labeled with Nugent scores from top experts. A separate set of 5815 images were evaluated by the CNN models. The best CNN model was selected to generalize its application on an independent sets of 1082 images collecting from three teaching hospitals. Different hardwares were used to take images in hospitals. ResultsOur model could classify three Nugent Scores from images with high three classification accuracy of 89.3% (with 82.4% sensitivity and 96.6% specificity) on the 5815 test images, which was better diagnostic yield than the top-level technologists and obstetricians in China. The ability of generalization for our model was strong that it obtained 75.1%, which was 6.6% higher than the average of technologists. ConclusionThe CNN model over performed human healthcare practitioners on accuracy, efficiency and stability for BV diagnosis using microscopic image-based Nugent scores. The deep learning model may offer translational application in automating diagnosis of bacterial vaginosis with proper supporting hardware.

microbiology↗

bioRxiv2020

Loss of FSTL1-expressing adipocyte progenitors drives the age-related involution of brown adipose tissue

In humans, brown adipose tissue (BAT) undergoes progressive involution or atrophy with increasing age, as manifested by decreased prevalence and mass, transformation to white adipose tissue (WAT), and reduction in thermogenic activity. This involution process cannot be fully recapitulated in rodent models and thus underlying cellular mechanisms are poorly understood. Here, we show that the interscapular BAT (iBAT) in rabbits involutes rapidly in early life, similarly to that in humans. The transcriptomic remodeling and identity switch of mature adipocytes are accompanied with the loss of brown adipogenic competence of their precursor cells. Through single-cell RNA sequencing, we surveyed the heterogenous populations of mesenchymal cells within the stromal vascular fraction of rabbit and human iBAT. An analogous FSTL1high population of brown adipocyte progenitors exists in both species while gradually disappear during iBAT involution in rabbits. In mice, FSTL1 is highly expressed by adipocyte progenitors in iBAT and genetic deletion of FSTL1 causes defective WNT signaling and iBAT atrophy in neonates. Our results underscore the BAT-intrinsic contribution from FSTL1high progenitors to age-related tissue involution and point to a potential therapeutic approach for obesity and its comorbidities. HIGHLIGHTSO_LIRabbit BAT irreversibly transforms to WAT before puberty. C_LIO_LIiBAT adipocyte progenitors reprogram transcriptome and lose brown adipogenic ability. C_LIO_LIComparable FSTL1high brown adipocyte progenitors exist in rabbit and human iBAT. C_LIO_LILoss of FSTL1 in brown adipocyte progenitors causes iBAT atrophy in mice. C_LI

physiology↗

bioRxiv2020

Individual Resting-state alpha peak frequency and within-trial changes in alpha peak frequency both predict visual flash segregation performance

Although sensory input is continuous, information must be combined over time to guide action and cognition, leading to the proposal of temporal sampling windows. A number of studies have suggested that a 10Hz sampling window might be involved in the "frame rate" of visual processing. To investigate this, we tested the ability of participants to localize and enumerate 1 or 2 visual flashes presented either at near-threshold (NT) or full contrast intensities, while recording magnetoencephalography (MEG). Performance was linked to the alpha frequency both at the individual level and trial-by-trial. Participants with a higher resting state alpha peak frequency showed the greatest improvement in performance as a function of ISI within the 100ms time window, while those with slower alpha improved more when ISI exceeded 100ms. On each trial, correct enumeration (1 vs. 2) performance was paired with faster pre-stimulus instantaneous alpha frequency. The effect of the timing of the NT stimulus on the perception was consistent with visual temporal integration windows being embedded in a sampling rhythm. Our results suggest that visual sampling/processing speed, linked to peak alpha frequency, is both an individual trait and varies in a state-dependent manner. Significance StatementA fundamental question in sensory and cognitive neuroscience is how the brain makes sense of the continuous flow of sensory input, parsing it into meaningful objects and events. The speed of cortical alpha rhythms is hypothesized to predict the temporal resolution of visual perception. We present a magnetoencephalography study investigating whether this temporal resolution is an individual trait or, in contrast, depends on fluctuations in brain state. Our results show that both individual resting state alpha frequency, a relatively stable trait, and trial-by-trial fluctuations of the instantaneous alpha frequency determine temporal segregation performance. These results have important implications for how our moment-by-moment perceptual experience is shaped as well as future intervention strategies to improve visual processing for specific tasks.

neuroscience↗

bioRxiv2020

Comparative transcriptome analysis reveals the intensive early-stage responses of host cells to SARS-CoV-2 infection

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a widespread outbreak of highly pathogenic COVID-19. It is therefore important and timely to characterize interactions between the virus and host cell at the molecular level to understand its disease pathogenesis. To gain insights, we performed high-throughput sequencing that generated time-series data simultaneously for bioinformatics analysis of virus genomes and host transcriptomes implicated in SARS-CoV-2 infection. Our analysis results showed that the rapid growth of the virus was accompanied by an early intensive response of host genes. We also systematically compared the molecular footprints of the host cells in response to SARS-CoV-2, SARS-CoV and MERS-CoV. Upon infection, SARS-CoV-2 induced hundreds of up-regulated host genes hallmarked by a significant cytokine production followed by virus-specific host antiviral responses. While the cytokine and antiviral responses triggered by SARS-CoV and MERS-CoV were only observed during the late stage of infection, the host antiviral responses during the SARS-CoV-2 infection were gradually enhanced lagging behind the production of cytokine. The early rapid host responses were potentially attributed to the high efficiency of SARS-CoV-2 entry into host cells, underscored by evidence of a remarkably up-regulated gene expression of TPRMSS2 soon after infection. Taken together, our findings provide novel molecular insights into the mechanisms underlying the infectivity and pathogenicity of SARS-CoV-2.

microbiology↗

bioRxiv2020

Structure of the ForT/PRPP complex uncovers the mechanism of C-C bond formation in C-nucleotide antibiotic biosynthesis

C-C bond formation is at the heart of anabolism and organic chemistry, but relatively few enzymatic strategies for catalyzing this reaction are known. The enzyme ForT catalyzes C-C bond formation between 5-phosphoribosyl-1-pyrophosphate (PRPP) and 4-amino-1H-pyrazole-3,5-dicarboxylate to make a key intermediate in the biosynthesis of the C-nucleotide formycin A 5-phosphate; we now report the 2.5 [A] resolution structure of the ForT/PRPP complex and thus locate the active site. Site-directed mutagenesis has identified those residues critical for PRPP recognition and catalysis. Structural conservation with GHMP kinases suggests that stabilization of the negatively charged pyrophosphate leaving group is crucial for catalysis in ForT. A mechanism for this new class of C-C bond forming enzymes is proposed. Entry for the Table of Contents O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=180 SRC="FIGDIR/small/009662v1_ufig1.gif" ALT="Figure 1"> View larger version (12K): org.highwire.dtl.DTLVardef@1817d54org.highwire.dtl.DTLVardef@4b3558org.highwire.dtl.DTLVardef@19b248eorg.highwire.dtl.DTLVardef@7d0c62_HPS_FORMAT_FIGEXP M_FIG C_FIG A new class of enzymes catalyse C-C bond formation by irreversible CO2 and pyrophosphate production.

biochemistry↗

bioRxiv2020

Preserving Vascular Integrity Protects Mice Against Multidrug-Resistant Gram-Negative Bacterial Infection

The rise in multidrug resistant (MDR) organisms portends a serious global threat to the healthcare system with nearly untreatable infectious diseases, including pneumonia and its often fatal sequelae, acute respiratory distress syndrome (ARDS) and sepsis. Gram-negative bacteria (GNB) including Acinetobacter baumannii, Pseudomonas aeruginosa, and carbapenemase-producing Klebsiella pneumoniae (CPKP), are among the World Health Organization and National Institutes of Healths high priority MDR pathogens for targeted development of new therapies. Here we show that stabilizing the hosts vasculature by genetic deletion or pharmacological inhibition of the small GTPase ADP-ribosylation factor 6 (ARF6) increases survival rates of mice infected with A. baumannii, P. aeruginosa, CPKP pneumonia. We show that pharmacological inhibition of ARF6-GTP phenocopies endothelial-specific Arf6 disruption in enhancing survival of mice with A. baumannii pneumonia, suggesting that inhibition is on target. Finally, we show that the mechanism of protection elicited by these small molecule inhibitors is by restoration of vascular integrity disrupted by GNB lipopolysaccharide (LPS) activation of TLR4/MyD88/ARNO/ARF6 pathway. By targeting the hosts vasculature with small molecule inhibitors of ARF6 activation, we circumvent microbial drug resistance and provide a potential alternative/adjunctive treatment for emerging and re-emerging pathogens.

microbiology↗

bioRxiv2020

Expression and purification of p72 trimers as subunit vaccine candidate

The authors have withdrawn this manuscript because the result is invalid. We intend to revise it on our technical approach and interpretation of the results. The authors do not wish for this manuscript to be cited as a reference for this project. Please contact the corresponding author if you have any questions.

microbiology↗

bioRxiv2020

Nelfinavir was predicted to be a potential inhibitor of 2019 nCov main protease by an integrative approach combining homology modelling, molecular docking and binding free energy calculation

2019-nCov has caused more than 80 deaths as of 27 January 2020 in China, and infection cases have been reported in more than 10 countries. However, there is no approved drug to treat the disease. 2019-nCov Mpro is a potential drug target to combat the virus. We built homology models based on SARS Mpro structures, and docked 1903 small molecule drugs to the models. Based on the docking score and the 3D similarity of the binding mode to the known Mpro ligands, 4 drugs were selected for binding free energy calculations. Both MM/GBSA and SIE methods voted for nelfinavir, with the binding free energy of -24.69{+/-}0.52 kcal/mol and -9.42{+/-}0.04 kcal/mol, respectively. Therefore, we suggested that nelfinavir might be a potential inhibitor against 2019-nCov Mpro.

pharmacology and toxicology↗

bioRxiv2019

Collaborative Behavior of Urea and KI in Denaturing Protein Native Structure

In this work, the combined influence of urea and KI on protein native structure is quantitatively investigated through the comparative molecular dynamics simulations on the structural dynamics of a polypeptide of TRPZIP4 in a series of urea/KI mixed solutions (urea concentration: 4M, KI salt concentration: 0M-6M). The observed enhanced denaturing ability of urea/KI mixture can be explained by direct interactions of urea/K+/water towards protein (electrostatic and vdW interactions from urea and electrostatic interactions from K+ and water) and indirect influence of KI on the strengthened interaction of urea towards protein backbone and side-chain. The latter indirect influence is fulfilled through the weakening of hydrogen bonding network among urea and water by the appearance of K+-water and I--urea interactions. As a result, the denaturing ability enhancement of urea and KI mixed solution is induced by the collaborative behavior of urea and KI salt.

biophysics↗