Biology subjects

Wang, J.

Publications and source records attributed to Wang, J..

At least 685 records · Page 38Linked to original sources

bioRxiv2025

Deepening imaging-based spatial proteomics at high spatial resolution through controlled tissue resizing and in-situ bottom-up mass spectrometry

Understanding the spatial organization of proteins in tissues is essential for elucidating biological function, but current proteomic imaging methods face limitations in resolution, sensitivity, and multiplexing capability. Here, we present IMPACT (Imaging Mass Spectrometry for Proteome Analysis on Compressed Tissue), a hydrogel-based workflow that integrates controlled vertical tissue compression following expansion to enhance protein detection while improving spatial resolution in mass spectrometry imaging (MSI). Through a 6-fold lateral expansion and 100-fold thickness reduction, analytes were concentrated at the surface to improve detection and enable analysis of thick specimen. Coupled with optimized in-gel digestion using hybrid enzymes, IMPACT increased protein identifications 3-fold compared to conventional MSI while maintaining spatial fidelity. We demonstrate IMPACTs utility in mapping low-abundance proteins and resolving fine anatomical structures. Orthogonal validation with immunofluorescence and integration with spatial transcriptomics further reveal protein-RNA distribution discrepancies, highlighting the need for direct proteomic imaging. IMPACT bridges critical gaps in spatial proteomics by enabling high-resolution, high-sensitivity, and multiplexed untargeted protein imaging, opening new avenues for tissue biology and biomarker discovery.

biophysics↗

bioRxiv2025

In vivo AAV9-SB-CRISPR screen identifies fatty acid elongase ELOVL5 as a pro-resolving mediator in lung inflammation

Timely resolution of inflammation is essential to prevent tissue damage and maintain homeostasis. Immunometabolism is critical for innate immunity and inflammation. However, how metabolic enzymes and metabolites contribute to inflammatory resolution remains largely unknown. To identify the key metabolic mediators of inflammation resolution, we generated an AAV9-Sleeping Beauty CRISPR library comprising 17090 sgRNAs targeting 2682 mouse metabolic genes. We then conducted an in vivo CRISPR screen in type II alveolar epithelial cells (AECIIs)-specifically expressing Cas9 mice and uncovered a very long chain fatty acid elongase, ELOVL5, that promoted the resolution of lung inflammation after influenza virus infection. Deficiency of Elovl5 in mouse lung epithelial cells impaired lung inflammation resolution and tissue repair phenotype both in vitro and in vivo. Mechanistically, ELOVL5 bound to STING, inhibiting TBK1 interaction and translocation to the Golgi. These effects ultimately reduced STING-mediated inflammation and promoted AKT1-mediated tissue repair. In addition, ELOVL5 decreased eicosanoid levels in AECIIs to promote lung inflammation resolution. Supplement with ELOVL5 downstream products reversed the increased expression of inflammatory cytokines caused by Elovl5 deficiency. These results support an unrevealed mechanism for polyunsaturated fatty acid metabolism in the resolution of innate inflammation and provide paths toward treating inflammatory diseases through manipulating cellular lipid metabolism.

immunology↗

bioRxiv2025

Systematic characterization of the ovarian landscape across mouse menopause models

Menopause affects not only fertility but also systemic health. Yet mechanisms underlying this coupling are still poorly understood, partly due to the absence of robust, age-relevant preclinical models with comprehensive molecular and phenotypic characterization. To address this, we systematically compared three candidate mouse models of menopause: (1) intact aging, (2) chemical ovarian follicle depletion using 4-vinylcyclohexene diepoxide (VCD) administered at multiple ages, and (3) Foxl2 haploinsufficiency, a genetic model based on a gene linked to human premature ovarian failure. Through histology, serum hormone profiling, single-cell transcriptomics and machine-learning approaches, we identify both shared and model-specific features of follicle loss, endocrine disruption, and transcriptional remodeling. Our comparative framework enables informed selection of context-appropriate preclinical rodent models to study menopause and the broader physiological consequences of ovarian aging. TeaserThis systematic characterization of mouse menopause models lays the groundwork for future preclinical work on the systemic impacts of menopause.

genomics↗

bioRxiv2025

rAAV-miniBEND: A targeted vector for brain endothelial cell gene delivery and cerebrovascular malformation modeling

Defects in brain endothelial cells (brainECs) can cause severe cerebrovascular malformations, including arteriovenous malformation (AVM) and cerebral cavernous malformation (CCM). The lack of appropriate tools for cerebrovascular disease modeling and local genetic manipulation of the brain vasculature hinders research on cerebrovascular malformations. Here we develop a recombinant adeno-associated virus (rAAV) tool miniBEND (rAAV-based mini-system for brain endothelial cells, rAAV-miniBEND), which combines a minimal promoter and an optimized cis-acting element (cis-element) isolated from the mouse gene Tek. This system achieves gene expression specifically in mouse and rat brainECs. Using rAAV-miniBEND, we achieve high-efficiency and high-specificity gene expression in brainECs through intracranial injection at various developmental stages and through intravenous administration at all postnatal stages in mice. Furthermore, we use rAAV-miniBEND to model sporadic CCMs mediated by MAP3K3I441M and AVMs mediated by BrafV600E. We demonstrate that somatic expression of BrafV600E in brainECs induces an AVM phenotype, and that brainEC proliferation are important for AVM development. Thus, our rAAV-miniBEND system provides a valuable and widely applicable tool for cerebrovascular disease modeling and local or global brainEC gene delivery.

neuroscience↗

bioRxiv2025

Highly accurate protein structure prediction-based virtual docking pipeline accelerating the identification of anti-schistosomal compounds

Schistosomiasis is a major neglected tropical disease that lacks an effective vaccine and faces increasing challenges from praziquantel resistance, underscoring the urgent need for novel therapeutics. Target-based drug discovery (TBDD) is a powerful strategy for drug development. In this study, we utilized AlphaFold to predict the structures of target proteins from Schistosoma mansoni and Schistosoma japonicum, followed by virtual molecular screening to identify potential inhibitors. Among 202 potential therapeutic targets, we identified 37 proteins with high-accuracy structural predictions suitable for molecular docking with 14,600 compounds. This screening yielded 268 candidate compounds, which were further evaluated in vitro for activity against both adult and juvenile S. mansoni and S. japonicum. Seven compounds exhibited strong anti-schistosomal activity, with HY-B2171A (Carubicin hydrochloride, CH) emerging as the most potent. CH was predicted to target the splicing factor U2AF65, and knockdown of its coding gene Smp_019690 resulted in a phenotype similar to CH treatment. RNA sequencing revealed that both CH treatment and Smp_019690 RNAi disrupted splicing events in the parasites. Further studies demonstrated that CH impairs parasite viability by inhibiting U2AF65 function in mRNA splicing regulation. By integrating RNAi-based target identification with structure-based virtual screening, alongside in vitro phenotypic and molecular analyses of compound-treated schistosomes, our study provides a comprehensive framework for anti-schistosomal drug discovery and identifies promising candidates for further preclinical development. Author SummarySchistosomiasis is a debilitating disease caused by parasitic worms and affects over 230 million people worldwide. For decades, control of the disease has relied on a single drug, praziquantel, which is less effective against juvenile parasites and increasingly threatened by the risk of drug resistance. To accelerate the discovery of new treatments, we integrated advanced computational tools with molecular validation. Using AlphaFold-predicted structures of 37 key schistosome proteins, we performed large-scale virtual screening to identify promising drug candidates. Over 200 compounds were prioritized for phenotypic screening on two major human-infective schistosome species, leading to the discovery of seven with potent anti-parasitic activity. Among them, Carubicin hydrochloride showed strong efficacy by targeting a critical splicing factor, U2AF65. Inhibiting this protein--either chemically or through gene silencing-- disrupted parasite movement, suppressed cell proliferation, and altered splicing events. Our work highlights the power of integrating AI-based protein modeling with phenotypic and mechanistic validation to accelerate drug discovery for neglected tropical diseases like schistosomiasis.

pathology↗

bioRxiv2025

HSV-1 orchestrates host RAP80 ubiquitination by ICP0 and UL36USP to promote viral survival

Herpes simplex virus 1 (HSV-1) is a widespread human pathogen that establishes lifelong infections, ubiquitination is crucial for HSV - 1 to interact with the host and regulate its life activities. But the mechanisms of how HSV-1 dynamically modulates host proteins at different infection stages to promote its own survival remains challenging. Here we identified DNA damage response protein RAP80 as a critical target of HSV-1 ubiquitination enzymes ICP0 and UL36USP throughout HSV-1s infection cycle. In early stages, RAP80 inhibits HSV-1 transcription by directly binding to the viral genome, blocking viral protein ICP4 binding to transcription factors. This interaction is regulated by phase separation via an intricate interplay between RAP80 and ICP0/ICP4. As infection progresses, the viral E3 ligase ICP0 degrades RAP80, dissolving phase separation, and allowing the formation of a mature viral replication compartment. In late infection stages, RAP80 is deubiquitinated by the viral deubiquitinase UL36USP, which restores cellular homeostasis and promotes HSV-1 survival. This process involves modulating the R-loop-cGAS-apoptosis pathway. These findings underscore the dynamic interplay between viral and host factors and the complex mechanisms used by HSV-1 to subvert host defenses, offering practical implications for the future development of antiviral strategies.

molecular biology↗

bioRxiv2025

Multiphasic Organization and Differential Dynamics of Proteins Within Protein-DNA Biomolecular Condensates

Biomolecular condensates formed through liquid-liquid phase separation are increasingly recognized as critical regulators of genome organization and gene expression. While the role of proteins in driving phase separation is well-established, how DNA modulates the structure and dynamics of protein-DNA condensates remains less well understood. Here, we employ a minimalist coarse-grained model to investigate the interplay between homotypic protein-protein and heterotypic protein-DNA interactions in governing condensate formation, composition, and internal dynamics. Our simulations reveal that DNA chain length and flexibility critically influence condensate morphology, leading to the emergence of multiphasic and core-shell organizations under strong heterotypic interactions. We find that DNA recruitment into the condensate significantly alters protein mobility, giving rise to differential dynamics of proteins within the condensate. By analyzing the distribution profiles of protein displacements, we identify up to five distinct diffusion modes, including proteins bound to DNA, confined within the dense phase, or freely diffusing. These results provide a mechanistic framework for interpreting spatially heterogeneous protein dynamics observed in chromatin condensates and emphasize the direct role of DNA in tuning condensate properties. Our findings provide new insights into how biophysical parameters may control the functional architecture of protein-DNA condensates in biological systems. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=158 SRC="FIGDIR/small/658691v1_ufig1.gif" ALT="Figure 1"> View larger version (73K): org.highwire.dtl.DTLVardef@108a818org.highwire.dtl.DTLVardef@153f9a1org.highwire.dtl.DTLVardef@29f220org.highwire.dtl.DTLVardef@1d74d6e_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2025

Controlling Multicomponent Condensate Morphology via Additive-Modulated Interactions

The morphology of biomolecular condensates plays a critical role in regulating intracellular organization and function by enabling both spatial and temporal control over biochemical processes. Recent studies have shown that small-molecule cosolutes can not only modulate phase separation but also influence condensate morphology. However, the mechanistic understanding of how small molecules regulate condensate structure remains limited. In this study, we employ coarse-grained molecular dynamics simulations to investigate how the morphology of two-component condensates can be modulated through the introduction of additional small-molecule cosolutes. By systematically varying the interaction strengths between the small molecules and the macromolecular components, we observe morphological transitions between, e.g., core-shell and dewetted structures. To rationalize these transitions, we calculate second virial coefficients in the presence of the small molecules, providing a molecular-level framework to capture shifts in effective homotypic and heterotypic interactions. We further investigate the role of stoichiometry between the small molecules and macromolecules, demonstrating that stoichiometry and interaction strength jointly determine the condensate morphology by altering the relative interaction strengths among components. Additionally, we show that fully miscible two-component condensates can undergo transitions to microphase-separated morphologies, such as core-shell or dewetted, upon small molecule introduction. Together, these findings reveal that condensate morphology can be rationally tuned through interaction- and stoichiometry-dependent mechanisms, offering molecular-scale insights into how small-molecule cosolutes modulate condensate structure.

biophysics↗

bioRxiv2025

Beyond the G-Loop: CRBN Molecular Glues Potently Target VAV1 via a Novel SH3 RT-Loop Degron

This study reports the discovery and characterization of novel CRBN molecular glues that selectively induce the proteasomal degradation of the hematopoietic-specific signaling protein VAV1, a key target in hematological malignancies and autoimmune diseases. Utilizing unbiased global proteomics, we identified phenyl-glutarimide derivatives NGT-201-12, as effective VAV1 degraders, with its C-terminal SH3 domain (SH3-2) being crucial for this interaction. A significant finding is the elucidation of a non-canonical RT-loop degron (RDxS motif, residues 796-799) within VAV1 SH3-2, distinct from previously characterized G-loop degrons. This discovery, supported by advanced computational modeling using the physics- and AI-driven GluePlex workflow and validated by site-directed mutagenesis, highlights versatility of CRBN in recognizing diverse neosubstrate motifs. Furthermore, we demonstrate that applying Free Energy Perturbation (FEP+) calculations to these predicted ternary structures yields cooperativity metrics that correlate with experimental degradation potency, overcoming the limitations of standard molecular docking. This establishes a robust workflow where, once a ternary complex is predicted, even with initial weak binders, FEP+ can be utilized to prospectively rank analogs and optimize molecular glue potency. Additionally, we demonstrate that strategic chemical modifications, particularly conformational restriction via halogen substitution (e.g., NGT-201-18), markedly potentiate VAV1 degradation, a principle supported by density functional theory (DFT) calculations. Comprehensive structure-activity relationship (SAR) studies provided a roadmap for designing next-generation VAV1 degraders. Importantly, dose-response proteomics not only confirmed VAV1 as the primary target but also revealed LIMD1, possessing a canonical G-loop, as an off-target for some analogs, indicating a single molecular glue can engage disparate degron motifs. The identification of the VAV1 RT-loop degron prompted a proteome-wide search, revealing other SH3-containing proteins as potential targets or off-targets. In conclusion, this research unveils a novel non-canonical RT-loop degron in VAV1, demonstrates the utility of conformational restriction in enhancing degrader potency, and underscores the critical role of integrating global proteomics with advanced structural modeling and FEP calculations for understanding degrader potency and selectivity. These findings offer a promising therapeutic strategy for targeting VAV1 and significantly expand the landscape of CRBN neosubstrate recognition and the rational design of molecular glue degraders.

biochemistry↗

bioRxiv2025

Engineering gene expression dynamics via self-amplifying RNA with drug-responsive non-structural proteins

The design of gene therapies with drug-regulatable expression of therapeutic payloads is of interest for diverse applications. We hypothesized that a regulated expression system based on alphavirus-derived self-amplifying RNAs (saRNAs), which encode 4 non-structural proteins (nsPs) that copy the RNA backbone to enable sustained expression, would have advantages in safety and simplicity of delivery. Here we designed saRNAs where payload expression is regulated by the FDA-approved drug trimethoprim (TMP), by fusing TMP-responsive degradation domains (DDs) to nsPs to regulate RNA self-amplification. Screening a library of nsP-DD fusions, we identified an optimal design with DDs fused to nsP2, nsP3, and the payload, achieving a high fold-change in expression level in response to TMP and low expression in the off state. In mice, this saRNA circuit enabled diverse dynamic expression patterns in response to oral TMP. Implementing this circuit for controlled expression of an HIV antigen, an escalating TMP regimen significantly enhanced germinal center responses critical for B cell affinity maturation. This drug- regulated RNA technology holds potential for vaccines, immunotherapies, and gene therapies.

bioengineering↗

bioRxiv2025

Inhibiting RhoA Activation via GDP-state Stabilization to Relieve Heart Failure

High rates of heart failure (HF) morbidity and mortality have made targeting myocardial remodeling--particularly hypertrophy and fibrosis--a key therapeutic focus. RhoA, which regulates cytoskeletal reorganization and cell migration, plays a role in this process. However, RhoA has long been considered "undruggable", due to its strong binding to its endogenous substrates, GDP/GTP, and the lack of well-defined pockets for drug targeting. Here, we discovered a cryptic pocket proximate to GDP within RhoA and identified a natural product, AH001, binds here and interacts with GDP, stabilizing RhoAs interaction with its endogenous inhibitor, RhoGDI. AH001 reduced the downstream MRTFA nuclear translocation and downregulated fibrosis/hypertrophy proteins. Consequently, AH001 mitigated myocardial remodeling in multiple HF animal models, and in the 3D myocardial tissue model. Our findings highlight the therapeutic potential of inhibiting RhoA activation in myocardial remodeling, ultimately targeting HF, and offer a promising avenue for developing reversible inhibitors against undruggable GTPases.

pharmacology and toxicology↗

bioRxiv2025

rtmsEcho: An Open-Source R Package for Automated Analysis of Acoustic Ejection Mass Spectrometry Data

Mass spectrometry (MS) is a well-established technology in biological research, enabling sensitive and precise quantitative analysis of complex samples. While traditional LC-MS systems provide robust performance for targeted analyses, their reliance on chromatographic separation limits throughput, rendering large-scale studies inefficient. The emergence of acoustic ejection mass spectrometry (AEMS) has revolutionized high-throughput workflows by eliminating chromatography and enabling direct nanoliter-scale sampling, achieving hundreds to thousands of measurements per hour. However, AEMSs full potential remains constrained by software limitations--existing tools lack robust automated processing capabilities for critical tasks such as peak detection, integration, and multimodal data analysis (e.g., multiple reaction monitoring, precursor ion, and neutral loss scans). To address this gap, we developed rtmsEcho, an open-source R package that extends our previously published rtms framework. This specialized solution provides direct access to AEMS data, enabling customizable processing of both MRM and full-scan acquisitions (precursor ion and neutral loss modes) while automating shot-to-peak association and spectral analysis. By streamlining data extraction and quantification, rtmsEcho enhances efficiency and reproducibility in high-throughput applications, including drug discovery, quality control, and clinical diagnostics. This innovation bridges a critical gap in AEMS data analysis, allowing researchers to fully leverage the speed and precision of next-generation mass spectrometry.

biochemistry↗

bioRxiv2025

H3K4me1 directs H3K36me2 and H3K36me3 deposition in land plants

Monomethylation of histone H3 lysine 4 (H3K4me1) marks enhancers in mammals. However, the function of H3K4me1 in plants remains largely unclear. Here, we analyzed the genome-wide distribution of H3K4me1 in diverse species across evolution, revealing a distinctive H3K4me1 distribution pattern in land plants. To explore the function of H3K4me1 in plants, we identified an H3K4me1-specific reader protein, Early heading date 3 (Ehd3), and solved the structure of Ehd3 in complex with the H3K4me1 peptide, revealing a unique binding module differing from the previously reported PHD finger proteins. We further identified an Ehd3-binding protein, SET domain group 724 (SDG724), and the deletion of either Ehd3 or SDG724 caused similar defects in plant phenotype and changes in transcriptome and epigenome profiles. Both Ehd3 and SDG724 are enriched at chromatin regions marked by H3K4me1 but not H3K4me2 or H3K4me3. Ehd3 activates the H3K36 methyltransferase SDG724, and H3K36me2/me3 are colocalized with H3K4me1 in the genomes of land plants. Collectively, our results reveal that H3K4me1 directs the establishment of H3K36me2 and H3K36me3 in land plants.

molecular biology↗

bioRxiv2025

Quantitative Assessment of Protein-DNA Interactions via SYBR Green Fluorescence

Protein-DNA interactions are crucial for cellular processes, but current quantification methods lack sensitivity. We developed the Real-time PCR-based DNA Binding Assay (RP-DBA) to detect and quantify these interactions. The target protein, expressed as a Strep-tag II fusion, is purified and incubated with double-stranded DNA probes containing 4 bp 3 overhangs. Protein-DNA complexes are immobilized on Strep-Tactin beads, washed, and eluted. A complementary single-stranded DNA amplification arm is added, extended by Taq polymerase, and quantified via qPCR with SYBR Green. RP-DBA enables real-time kinetic analysis and is 4- to 10-fold more sensitive than EMSA, depending on amplification arm length (30-90 bp). Its simplicity, speed, accuracy, and high-throughput potential make it a valuable tool for advancing molecular biology research.

molecular biology↗

bioRxiv2025

bstinence and Extinction Drive Opposing Changes in Striatal Activity and Dopamine Signaling During Alcohol Relapse

Relapse remains a major obstacle in the treatment of alcohol use disorder, often driven in part by enduring neuroadaptations. However, how different treatment strategies--such as abstinence versus extinction training--modulate the underlying neural circuits and synaptic mechanisms that shape relapse vulnerability remains poorly understood. In this study, we demonstrate that abstinence and extinction distinctly influence dorsomedial striatal (DMS) direct-pathway medium spiny neuron (dMSN) activity and dopamine signaling during cue-induced reinstatement of alcohol seeking. Using in vivo fiber photometry in D1-Cre rats expressing calcium or dopamine sensors, we found that abstinence enhanced dMSN calcium responses and dopamine release during reinstatement, whereas extinction normalized these neural signals and suppressed relapse-like behavior. Furthermore, bidirectional optogenetic modulation of medial prefrontal cortex (mPFC)-to-dMSN synapses revealed a causal role for corticostriatal plasticity in determining relapse propensity. Inducing long-term depression (LTD) in the abstinent state attenuated reinstatement, while inducing long-term potentiation (LTP) after extinction training reinstated alcohol seeking. Together, these findings identify distinct neural adaptations shaped by abstinence versus extinction and highlight corticostriatal plasticity as a potential target for relapse prevention. HighlightO_LIAbstinence enhances striatal dMSN activity and dopamine signaling during cued relapse. C_LIO_LIExtinction training normalizes dMSN dynamics and reduces dopamine release during cued relapse. C_LIO_LIOptogenetic mPFC-to-dMSN long-term depression after abstinence reduces relapse. C_LIO_LIOptogenetic mPFC-to-dMSN long-term potentiation after extinction invigorates relapse. C_LI

neuroscience↗

bioRxiv2025

Phage intervention improves colitis and response to corticosteroids by attenuating virulence of Crohn's disease-associated bacteria

Adherent-invasive Escherichia coli (AIEC) exhibit proinflammatory properties and have been implicated in the pathogenesis of Crohns disease (CD), a form of inflammatory bowel disease (IBD). Antibiotic use in CD lacks specificity and may worsen microbiome disruption, prompting interest in bacteriophages (phages) for targeted microbiome editing. Here, we identified HER259, a phage active against the clinical AIEC strain NRG857c. Using gnotobiotic models of AIEC-driven colitis, we show that HER259 attenuates AIEC virulence, including suppression of the FimH adhesin through inversion of the fimS promoter to its off orientation. Withdrawal of HER259 treatment leads to reversion of the fimS promoter and reactivated colitis in mice. HER259 phage also enhances the therapeutic effect of sub-therapeutic budesonide, independent of microbial drug metabolism. These findings support targeted phage therapy as an adjunct treatment approach in IBD, demonstrating modulation of bacterial virulence and improved response to conventional treatments which may reduce drug-related side effects. One Sentence SummaryBacteriophage HER259 improves colitis severity mediated by Crohns disease Escherichia coli NRG857c, and increases efficacy of budesonide.

microbiology↗

bioRxiv2025

Elucidating the mechanisms of action and evolutionary history of phage anti-defence proteins

Phages and bacteria are locked in a molecular arms race, with phage anti-defence proteins (ADPs) enabling them to evade bacterial immune systems. To streamline access to information on ADPs, we developed the Encyclopaedia of Viral Anti-DefencE Systems (EVADES), an online resource containing sequences, structures, protein family annotations, and mechanisms of action (MoA) for 268 ADPs. Through computational structural analysis we predicted MoAs for 21 uncharacterised ADPs. We demonstrate the utility of EVADES by exploring different characteristics of ADPs: (i) DNA mimic ADPs exhibit broad-spectrum activity against bacterial defence systems; (ii) protein sharing across defence systems enables multi-defence inhibitory activity of ADPs; and (iii) eukaryotic dsDNA viruses encode ADP homologs, suggesting conserved immune evasion strategies across domains of life. These results highlight the broader relevance of phage ADPs in understanding interactions between prokaryotic or eukaryotic viruses and their hosts. EVADES is freely accessible at https://www.ebi.ac.uk/finn-srv/evades. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=132 SRC="FIGDIR/small/658234v3_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@18460corg.highwire.dtl.DTLVardef@19ef231org.highwire.dtl.DTLVardef@2e91a5org.highwire.dtl.DTLVardef@15c2547_HPS_FORMAT_FIGEXP M_FIG C_FIG O_LIThe EVADES database aggregates knowledge on phage anti-defence proteins (ADPs), facilitating hypothesis generation. C_LIO_LIMechanisms of action are proposed for 21 uncharacterised ADPs. C_LIO_LIADPs that function as DNA mimics can block diverse bacterial defence systems. C_LIO_LIHomology of targets of ADPs underpin their broad counter-defence capacities and enable prediction of further targets. C_LIO_LIEukaryotic double-stranded DNA viruses employ functional homologs of ADPs to evade host immune responses. C_LI

microbiology↗

bioRxiv2025

Genome-wide CRISPR/Cas9 knockout screen identifies host factors essential for Bovine Parainfluenza Virus Type 3 replication

Bovine Parainfluenza Virus Type 3 (BPIV3) is a leading cause of respiratory illness in cattle and a primary component of Bovine Respiratory Disease Complex (BRDC), resulting in significant economic losses. Understanding the mechanisms of BPIV3 infection, particularly the entry process, is essential for developing effective control measures. Identifying specific host factors that viruses exploit during their life cycle can reveal critical vulnerabilities for potential antiviral targets. We established a genome-wide CRISPR/Cas9 knockout screen in bovine cells to identify host factors involved in viral infections. Our screen identified several key host factors required for BPIV3 infection, including the sialic acid transporter SLC35A1 and the protein LSM12. Further mechanistic analysis revealed that these factors play critical roles at distinct stages of the BPIV3 entry process. These findings not only advance our understanding of how BPIV3 infects host cells, but also identify potential host targets for inhibiting infection and developing novel antiviral strategies.

microbiology↗