Biology subjects

Xie, X.

Publications and source records attributed to Xie, X..

At least 235 records · Page 13Linked to original sources

bioRxiv2020

Natural food intake patterns do not synchronize peripheral clocks

Food is thought to synchronize circadian clocks in the body, but this is based on time-restricted feeding (TRF) protocols. To test whether naturalistic feeding patterns are sufficient to phase-shift and entrain peripheral tissues, we measured circadian rhythms of the liver, kidney, and submandibular gland in mPer2Luc mice under different feeding schedules. In ad lib feeding as well as in a schedule designed to mimic the ad lib pattern, PER2::LUC bioluminescence peaked during the night as expected. Surprisingly, shifting the scheduled feeding by 12h caused only small advances (<3h). To isolate the effects of feeding from the light-dark cycle, clock phase was then measured in mice acclimated to scheduled feeding and housed in constant darkness. In these conditions, peripheral clock phases were better predicted by the rest-activity cycle than the food schedule. Under natural feeding patterns, the master pacemaker in the brain sets the phase of peripheral organs independent of feeding behavior.

physiology↗

bioRxiv2020

A nanoluciferase SARS-CoV-2 for rapid neutralization testing and screening of anti-infective drugs for COVID-19

A high-throughput platform would greatly facilitate COVID-19 serological testing and antiviral screening. Here we report a nanoluciferase SARS-CoV-2 (SARS-CoV-2-Nluc) that is genetically stable and replicates similarly to the wild-type virus in cell culture. We demonstrate that the optimized reporter virus assay in Vero E6 cells can be used to measure neutralizing antibody activity in patient sera and produces results in concordance with a plaque reduction neutralization test (PRNT). Compared with the low-throughput PRNT (3 days), the SARS-CoV-2-Nluc assay has substantially shorter turnaround time (5 hours) with a high-throughput testing capacity. Thus, the assay can be readily deployed for large-scale vaccine evaluation and neutralizing antibody testing in humans. Additionally, we developed a high-throughput antiviral assay using SARS-CoV-2-Nluc infection of A549 cells expressing human ACE2 receptor (A549-hACE2). When tested against this reporter virus, remdesivir exhibited substantially more potent activity in A549-hACE2 cells compared to Vero E6 cells (EC50 0.115 vs 1.28 M), while this difference was not observed for chloroquine (EC50 1.32 vs 3.52 M), underscoring the importance of selecting appropriate cells for antiviral testing. Using the optimized SARS-CoV-2-Nluc assay, we evaluated a collection of approved and investigational antivirals and other anti-infective drugs. Nelfinavir, rupintrivir, and cobicistat were identified as the most selective inhibitors of SARS-CoV-2-Nluc (EC50 0.77 to 2.74 M). In contrast, most of the clinically approved antivirals, including tenofovir alafenamide, emtricitabine, sofosbuvir, ledipasvir, and velpatasvir were inactive at concentrations up to 10 M. Collectively, this high-throughput platform represents a reliable tool for rapid neutralization testing and antiviral screening for SARS-CoV-2.

microbiology↗

bioRxiv2020

β-actin dependent chromatin remodeling mediates compartment level changes in 3D genome architecture

{beta}-actin is a crucial component of several chromatin remodeling complexes that control chromatin structure and accessibility. The mammalian Brahma-associated factor (BAF) is one such complex that plays essential roles in development and differentiation by regulating the chromatin state of critical genes and opposing the repressive activity of polycomb repressive complexes (PRCs). While previous work has shown that {beta}-actin loss can lead to extensive changes in gene expression and heterochromatin organization, it is not known if changes in {beta}-actin levels can directly influence chromatin remodeling activities of BAF and polycomb proteins. Here we conduct a comprehensive genomic analysis of {beta}-actin knockout mouse embryonic fibroblasts (MEFs) using ATAC-Seq, HiC-seq, RNA-Seq and ChIP-Seq of various epigenetic marks. We demonstrate that {beta}-actin levels can affect the complex interplay between chromatin remodelers such as BAF/BRG1 and EZH2 in a dosage-dependent manner. Our results show that changes in {beta}-actin levels and associated chromatin remodeling activities can not only impact local chromatin accessibility but also induce reversible changes in 3D genome architecture. Our findings support a novel role for {beta}-actin-dependent chromatin remodeling in shaping the chromatin landscape and regulating genes involved in development and differentiation.

genomics↗

bioRxiv2020

Structural basis of bile acid receptor activation and Gs coupling

G protein-coupled bile acid receptor (GPBAR) is a membrane receptor that senses bile acids to regulate diverse functions through Gs activation. Here, we report the cryo-EM structures of GPBAR-Gs complexes stabilized by either high-affinity P395 or the semisynthesized bile acid derivative INT-777 at 3-[A] resolution. These structures revealed a large oval-shaped ligand pocket with several sporadic polar groups to accommodate the amphipathic cholic core of bile acids. A fingerprint of key residues recognizing diverse bile acids in the orthosteric site, a putative second bile acid binding site with allosteric properties and structural features contributing to bias property were identified through structural analysis and mutagenesis studies. Moreover, structural comparison of GPBAR with other GPCRs uncovered an atypical mode of receptor activation and G-protein- coupling, featuring a different set of key residues connecting the ligand binding pocket to the Gs coupling site, and a specific interaction motif localized in intracellular loop 3. Overall, our study not only provides unique structural features of GPBAR in bile acid recognition, allosteric effects and biased signaling, but also suggests that distinct allosteric connecting mechanisms between the ligand binding pocket and the G protein binding site exist in the GPCR superfamily.

biochemistry↗

bioRxiv2020

Broad-spectrum antivirals of protoporphyrins inhibit the entry of highly pathogenic emerging viruses

Severe emerging and re-emerging viral infections such as Lassa fever, Avian influenza (AI), and COVID-19 caused by SARS-CoV-2 urgently call for new strategies for the development of broad-spectrum antivirals targeting conserved components in the virus life cycle. Viral lipids are essential components, and viral-cell membrane fusion is the required entry step for most unrelated enveloped viruses. In this paper, we identified a porphyrin derivative of protoporphyrin IX (PPIX) that showed broad antiviral activities in vitro against a panel of enveloped pathogenic viruses including Lassa virus (LASV), Machupo virus (MACV), and SARS-CoV-2 as well as various subtypes of influenza A viral strains with IC50 values ranging from 0.91{+/-}0.25 M to 1.88{+/-}0.34 M. A mechanistic study using influenza A/Puerto Rico/8/34 (H1N1) as a testing strain showed that PPIX inhibits the infection in the early stage of virus entry through biophysically interacting with the hydrophobic lipids of enveloped virions, thereby inhibiting the formation of the negative curvature required for fusion and blocking the entry of enveloped viruses into host cells. In addition, the preliminary antiviral activities of PPIX were further assessed by testing mice infected with the influenza A/Puerto Rico/8/34 (H1N1) virus. The results showed that compared with the control group without drug treatment, the survival rate and mean survival time of the mice treated with PPIX were apparently prolonged. These data encourage us to conduct further investigations using PPIX as a lead compound for the rational design of lipid-targeting antivirals for the treatment of infection with enveloped viruses.

pharmacology and toxicology↗

bioRxiv2020

Transcriptional Difference between SARS-COV-2 and other Human Coronaviruses Revealed by Sub-genomic RNA Profiling

BackgroundFundamental to viral biology is identification and annotation of viral genes and their function. Determining the level of coronavirus gene expression is inherently difficult due to the positive stranded RNA genome and the identification of sub-genomic RNAs (sgRNAs) that are required for expression of most viral genes. In the COVID-19 epidemic so far, few genomic studies have looked at viral sgRNAs and none have systematically examined the sgRNA profiles of large numbers of SARS-CoV2 datasets in conjuction with data for other coronaviruses. ResultsWe developed a bioinformatic pipeline to analyze the sgRNA profiles of coronaviruses and applied it to 588 individual samples from 20 independent studies, covering more than 10 coronavirus species. Our result showed that SARS-CoV, SARS-CoV-2 and MERS-CoV each had a core sgRNA repertoire generated via a canonical mechanism. Novel sgRNAs that encode peptides with evolutionarily conserved structures were identified in several coronaviruses and were expressed in vitro and in vivo. Two novel peptides may have direct functional relevance to disease, by alluding interferon responses and disrupting IL17E (IL25) signaling. Relevant to coronavirus infectivity and transmission, we also observed that the level of Spike sgRNAs were significantly higher in-vivo than in-vitro, while the opposite held true for the Nucleocapside protein. ConclusionsOur results greatly expanded the predicted number of coronaviruses proteins and identified potential viral peptide suggested to be involved in viral virulence. These methods and findings shed new light on coronavirus biology and provides a valuable resource for future genomic studies of coronaviruses.

bioinformatics↗

bioRxiv2020

Microvascular Injury in Mild Traumatic Brain Injury Accelerates Alzheimer-like Pathogenesis in Mice

IntroductionTraumatic brain injury (TBI) is considered as the most robust environmental risk factor for Alzheimers disease (AD). Besides direct neuronal injury and neuroinflammation, vascular impairment is also a hallmark event of the pathological cascade after TBI. However, the vascular connection between TBI and subsequent AD pathogenesis remains underexplored. MethodsWe established a closed-head mild TBI (mTBI) model in mice with controlled cortical impact, and examined the time courses of microvascular injury, blood-brain barrier (BBB) dysfunction, gliosis and motor function impairment in wild type C57BL/6 mice. We also determined the brain clearance of {beta}-amyloid, as well as amyloid pathology and cognitive functions after mTBI in the 5xFAD mouse model of AD. ResultsmTBI induced microvascular injury with BBB breakdown, pericyte loss and cerebral blood flow reduction in mice, which preceded gliosis. mTBI also impaired brain amyloid clearance via the vascular pathways. More importantly, mTBI accelerated amyloid pathology and cognitive impairment in the 5xFAD mice. DiscussionOur data demonstrated that microvascular injury plays a key role in the pathogenesis of AD after mTBI. Therefore, restoring vascular functions might be beneficial for patients with mTBI, and potentially reduce the risk of developing AD.

neuroscience↗

bioRxiv2020

New Soluble Angiopoietin Analog of C4BP-ANG1 Prevents Pathological Vascular Leakage

Vascular leak is a key driver of organ injury in diseases such as Acute Respiratory Distress Syndrome caused by viruses, including COVID-19. Strategies that reduce enhanced permeability and vascular inflammation are promising therapeutic targets. Activation of the Angiopoietin-1 (Angpt1)-Tie2 tyrosine kinase signaling pathway is an important regulator of vascular quiescence. Here we describe the design and construction of a new soluble ANGPT1 mimetic that is a potent activator of endothelial Tie2 in vitro and in vivo. Using a chimeric fusion strategy, we replaced the extracellular matrix (ECM) binding and oligomerization domain of ANGPT1 with a heptameric scaffold derived from the C-terminus of serum complement protein C4-binding protein (C4BP). We refer to this new fusion protein biologic as C4BP-ANG1, which forms a stable heptamer and induces TIE2 phosphorylation in cultured cells, and in the lung following i.v. injection of mice. Injection of C4BP-ANG1 ameliorates VEGF- and lipopolysaccharide-induced vascular leakage, in keeping with the known functions of Angpt1-Tie2 in maintaining quiescent vascular stability, and therefore is a promising candidate treatment for inflammatory endothelial dysfunction.

biochemistry↗

bioRxiv2020

Structure-Based Design, Synthesis and Biological Evaluation of Peptidomimetic Aldehydes as a Novel Series of Antiviral Drug Candidates Targeting the SARS-CoV-2 Main Protease

SARS-CoV-2 is the etiological agent responsible for the COVID-19 outbreak in Wuhan. Specific antiviral drug are urgently needed to treat COVID-19 infections. The main protease (Mpro) of SARS-CoV-2 is a key CoV enzyme that plays a pivotal role in mediating viral replication and transcription, which makes it an attractive drug target. In an effort to rapidly discover lead compounds targeting Mpro, two compounds (11a and 11b) were designed and synthesized, both of which exhibited excellent inhibitory activity with an IC50 value of 0.05 M and 0.04 M respectively. Significantly, both compounds exhibited potent anti-SARS-CoV-2 infection activity in a cell-based assay with an EC50 value of 0.42 M and 0.33 M, respectively. The X-ray crystal structures of SARS-CoV-2 Mpro in complex with 11a and 11b were determined at 1.5 [A] resolution, respectively. The crystal structures showed that 11a and 11b are covalent inhibitors, the aldehyde groups of which are bound covalently to Cys145 of Mpro. Both compounds showed good PK properties in vivo, and 11a also exhibited low toxicity which is promising drug leads with clinical potential that merits further studies.

biochemistry↗

bioRxiv2020

Topoisomerase III-beta is required for efficient replication of positive-sense RNA viruses

Based on genome-scale loss-of-function screens we discovered that Topoisomerase III-{beta} (TOP3B), a human topoisomerase that acts on DNA and RNA, is required for yellow fever virus and dengue virus-2 replication. Remarkably, we found that TOP3B is required for efficient replication of all positive-sense-single stranded RNA viruses tested, including SARS-CoV-2. While there are no drugs that specifically inhibit this topoisomerase, we posit that TOP3B is an attractive anti-viral target.

microbiology↗

bioRxiv2020

Hydroxyl carlactone derivatives are predominant strigolactones in Arabidopsis

Strigolactones (SLs) regulate important aspects of plant growth and stress responses. Many diverse types of SL occur in plants, but a complete picture of biosynthesis remains unclear. In Arabidopsis thaliana, we have demonstrated that MAX1, a cytochrome P450 monooxygenase, converts carlactone (CL) into carlactonoic acid (CLA), and that LBO, a 2-oxoglutarate-dependent dioxygenase, converts methyl carlactonoate (MeCLA) into a metabolite called [MeCLA+16] Da. In the present study, feeding experiments with deuterated MeCLAs revealed that [MeCLA+16] Da is hydroxymethyl carlactonoate (1-HO-MeCLA). Importantly, this LBO metabolite was detected in plants. Interestingly, other related compounds, methyl 4-hydroxycarlactonoate (4-HO-MeCLA) and methyl 16-hydroxycarlactonoate (16-HO-MeCLA) were also found to accumulate in lbo mutants. 3-HO-, 4-HO- and 16-HO-CL were detected in plants, but their expected corresponding metabolites, HO-CLAs, were absent in max1 mutants. These results suggest that HO-CL derivatives are predominant SLs in Arabidopsis, produced through MAX1 and LBO.

plant biology↗

bioRxiv2020

A Simultaneous Methylome and Proteome Quantitative Workflow for Complex Samples

Proteoforms containing post-translational modifications (PTMs) represent a degree of functional diversity only harnessed through analytically precise simultaneous quantification of multiple PTMs. Here we present a method to accurately differentiate an unmodified peptide from its PTM-containing counterpart through data-independent acquisition-mass spectrometry, leveraging small precursor mass windows to physically separate modified peptidoforms from each other during MS2 acquisition. We utilize a lysine and arginine PTM-enriched peptide assay library and site localization algorithm to simultaneously localize and quantify seven PTMs including mono-, di-, and tri-methylation, acetylation, and succinylation in addition to total protein quantification in a single MS run without the need to enrich experimental samples. To evaluate biological relevance, this method was applied to liver lysate from differentially methylated non-alcoholic steatohepatitis (NASH) mouse models. We report altered methylation and acetylation together with total protein changes drive the novel hypothesis of a regulatory function of PTMs in protein synthesis and mRNA stability in NASH. Graphical Abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

systems biology↗

bioRxiv2020

Predicting transcription factor binding in single cells through deep learning

Characterizing genome-wide binding profiles of transcription factor (TF) is essential for understanding many biological processes. Although techniques have been developed to assess binding profiles within a population of cells, determining binding profiles at a single cell level remains elusive. Here we report scFAN (Single Cell Factor Analysis Network), a deep learning model that predicts genome-wide TF binding profiles in individual cells. scFAN is pre-trained on genome-wide bulk ATAC-seq, DNA sequence and ChIP-seq data, and utilizes single-cell ATAC-seq to predict TF binding in individual cells. We demonstrate the efficacy of scFAN by studying sequence motifs enriched within predicted binding peaks and investigating the effectiveness of predicted TF peaks for discovering cell types. We develop a new metric "TF activity score" to characterize each cell, and show that the activity scores can reliably capture cell identities. The method allows us to discover and study cellular identities and heterogeneity based on chromatin accessibility profiles.

bioinformatics↗

bioRxiv2020

Open syntaxin overcomes synaptic transmission defects in diverse C. elegans exocytosis mutants

Assembly of SNARE complexes that mediate neurotransmitter release requires opening of a closed conformation of UNC-64/syntaxin. Rescue of unc-13/Munc13 phenotypes by overexpressed open UNC-64/syntaxin suggested a specific function of UNC-13/Munc13 in opening UNC-64/ syntaxin. Here, we revisit the effects of open unc-64/syntaxin by generating knockin (KI) worms. The KI animals exhibited enhanced spontaneous and evoked exocytosis compared to wild-type animals. Unexpectedly, the open syntaxin KI partially suppressed exocytosis defects of various mutants, including snt-1/synaptotagmin, unc-2/P/Q/N-type Ca2+ channel alpha-subunit, and unc-31/CAPS in addition to unc-13/Munc13 and unc-10/RIM, and enhanced exocytosis in tom-1/Tomosyn mutants. However, open syntaxin aggravated the defects of unc-18/Munc18 mutants. Correspondingly, open syntaxin partially bypasses the requirement of Munc13 but not Munc18 for liposome fusion. Our results show that facilitating opening of syntaxin enhances exocytosis in a wide range of genetic backgrounds, and may provide a general means to enhance synaptic transmission in normal and disease states.

neuroscience↗

bioRxiv2019

Chemical identification of 18-hydroxycarlactonoic acid as an LjMAX1 product and conversion of 18-hydroxylcarlactonoates to canonical and non-canonical strigolactones in Lotus japonicus

Strigolactones (SLs) are a group of plant apocarotenoids that act as rhizosphere signaling molecules for both arbuscular mycorrhizal fungi and root parasitic plants. They also regulate plant architecture as phytohormones. The model legume Lotus japonicus produces canonical 5-deoxystrigol (5DS) and non-canonical lotuslactone (LL). The biosynthesis pathways of the two SLs remain elusive. In this study, we characterized the L. japonicus MAX1 homolog, LjMAX1, found in the Lotus japonicus genome assembly build 2.5. The L. japonicus max1 LORE1 insertion mutant was deficient in 5DS and LL production. A recombinant LjMAX1 protein expressed in yeast microsomes converted carlactone (CL) to 18-hydroxycarlactonoic acid (18-HO-CLA) via carlactonoic acid (CLA). Identity of 18-HO-CLA was confirmed by comparison of the methyl ester derivative of the MAX1 product with the chemically synthesized methyl 18-hydroycarlactonoate (18-HO-MeCLA) using LC-MS/MS. (11R)-CL was detected as an endogenous compound in the root of L. japonicus. 13C-labeled CL, CLA, and 18-HO-MeCLA were converted to [13C]-5DS and LL in plant feeding experiments using L. japonicus WT. These results showed that LjMAX1 is the crucial enzyme in the biosynthesis of Lotus SLs and that 18-hydroxylated carlactonoates are precursors for SL biosynthesis in L. japonicus.

plant biology↗

bioRxiv2019

In-Depth Characterization and Validation in BRG1-Mutant Lung Cancers Define Novel Catalytic Inhibitors of SWI/SNF Chromatin Remodeling

Members of the ATP-dependent SWI/SNF chromatin remodeling complexes are among the most frequently mutated genes in cancer, suggesting their dysregulation plays a critical role. The synthetic lethality between SWI/SNF catalytic subunits BRM/SMARCA2 and BRG1/SMARCA4 has instigated great interest in targeting BRM. Here we have performed a critical and in-depth investigation of novel dual inhibitors (BRM011 and BRM014) of BRM and BRG1 in order to validate their utility as chemical probes of SWI/SNF catalytic function, while obtaining insights into the therapeutic potential of SWI/SNF inhibition. In corroboration of on-target activity, we discovered compound resistant mutations through pooled screening of BRM variants in BRG1-mut cancer cells. Strikingly, genome-wide transcriptional and chromatin profiling (ATAC-Seq) provided further evidence of pharmacological perturbation of SWI/SNF chromatin remodeling as BRM011 treatment induced specific changes in chromatin accessibility and gene expression similar to genetic depletion of BRM. Finally, these compounds have the capacity to inhibit the growth of tumor-xenografts, yielding important insights into the feasibility of developing BRM/BRG1 ATPase inhibitors for the treatment of BRG1-mut lung cancers. Overall, our studies not only establish the feasibility of inhibiting SWI/SNF catalytic function, providing a framework for SWI/SNF therapeutic targeting, but have also yielded successful elucidation of small-molecule inhibitors that will be of importance in probing SWI/SNF function in various disease contexts.

cancer biology↗

bioRxiv2019

Single-cell transcriptome profiling reveals neutrophil heterogeneity and orchestrated maturation during homeostasis and bacterial infection

The full neutrophil heterogeneity and differentiation landscape remains incompletely characterized. Here we profiled >25,000 differentiating and mature mouse neutrophils using single-cell RNA sequencing to provide a comprehensive transcriptional landscape of neutrophil maturation, function, and fate decision in their steady state and during bacterial infection. Eight neutrophil populations were defined by distinct molecular signatures. The three mature peripheral blood neutrophil subsets arise from distinct maturing bone marrow neutrophil subsets. Driven by both known and uncharacterized transcription factors, neutrophils gradually acquire microbicidal capability as they traverse the transcriptional landscape, representing an evolved mechanism for fine-tuned regulation of an effective but balanced neutrophil response. Bacterial infection reprograms the genetic architecture of neutrophil populations, alters dynamic transition between each subpopulation, and primes neutrophils for augmented functionality without affecting overall heterogeneity. In summary, these data establish a reference model and general framework for studying neutrophil-related disease mechanisms, biomarkers, and therapeutic targets at single-cell resolution.\n\nGraphical Abstract\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=119 SRC=\"FIGDIR/small/792200v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (28K):\norg.highwire.dtl.DTLVardef@18fed35org.highwire.dtl.DTLVardef@1e68b7org.highwire.dtl.DTLVardef@6cfbc8org.highwire.dtl.DTLVardef@1061a26_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIA comprehensive single-cell resolution transcriptional landscape of mouse neutrophil maturation and fate decision under steady-state and bacterial infection conditions.\nC_LIO_LIThe pathogen clearance machinery in neutrophils is continuously and gradually built during neutrophil differentiation, maturation, and aging, driven by both known and uncharacterized transcription factors.\nC_LIO_LIThe three mature neutrophil subsets in peripheral blood, including a novel ISG-expressing subset, are derived from distinct bone marrow neutrophil precursors.\nC_LIO_LIBacterial infection reprograms the genetic architecture of neutrophil populations, alters dynamic transition between each subpopulation, and primes neutrophils for augmented functionality without affecting overall neutrophil heterogeneity.\nC_LIO_LIBacterial infection-induced emergency granulopoiesis is mediated by augmented proliferation of early stage neutrophil progenitors and accelerated post-mitotic maturation.\nC_LI

immunology↗

bioRxiv2019

Influencing factors of telephone-cardiopulmonary resuscitation in China: a qualitative exploration based on managerial perspectives

BackgroundTelephone-cardiopulmonary resuscitation(T-CPR) has been proven to systematically improve bystander CPR implementation and thus improve the survival rate of out-of-hospital cardiac arrest (OHCA) patients on a large scale. However, China has a lower proportion of cities that provide T-CPR than other countries.This study aimed to explore the factors affecting the providing of T-CPR based on managerial perspectives and promote the implementation of T-CPR in China to Protect human health. MethodsThis study adopted a descriptive qualitative method.The managers from health bureau and first-aid Center were recruited to participate through purposive sampling. Data were collected using semi-structured interviews and Colaizzi 7-step analysis method was adopted to summarize and conclude the theme. ResultsA total of 10 managers were interviewed.Five main themes were identified: (a) bystander factors, (b) dispatching factors, (c) legal factors, (d) guiding factors, and (e) financial factors. ConclusionIt is urgent to promote the implementation of T-CPR in China.We can promote it by strengthening the training of bystanders in CPR knowledge and skills, developing T-CPR guidance process suitable for Chines national conditions, building an intelligent prehospital emergency system, promoting the legislation of first aid exemption, and providing financial support from various channels.

systems biology↗