Biology subjects

Xiong, Y.

Publications and source records attributed to Xiong, Y..

At least 55 records · Page 3Linked to original sources

bioRxiv2026

Conserved assembly architecture of the essential herpesvirus packaging accessory factor

To create a new wave of infectious virions, all herpesviruses require an accessory factor of unknown function to package their viral genomes into nascent capsids. Here, we present cryo-EM structures of the packaging accessory factor from the -herpesvirus herpes simplex virus type 1 (HSV-1, UL32) and the {beta}-herpesvirus human cytomegalovirus (HCMV, UL52). Unlike homologs from the {gamma}-herpesviruses, neither UL32 nor UL52 form stable homopentameric rings. UL52 forms incomplete pentameric rings lacking one or two protomers. UL32 does not form stable higher-order species, but stabilization through chemical crosslinking revealed a novel quaternary structure where three pentameric rings assemble into a "tripentamer." Our results reveal that herpesvirus packaging accessory factors adopt distinct oligomeric states but are constrained to pentameric symmetry. Assembly of protomers into a ring creates a positively charged central channel that we show is critical for infectious virus production in HSV-1. Taken together, our study points to a structurally conserved, essential function of packaging accessory factors across the Herpesviridae. AUTHOR SUMMARYHerpesviruses are a diverse family of pathogens that drive human disease. At the end of their life cycle, all herpesviruses must actively package their genomes into newly formed capsids. Although some aspects of packaging are conserved with tailed bacteriophages, herpesviruses require an additional packaging accessory factor of unknown function. Here, we present two high-resolution structures of the packaging accessory factor from herpes simplex virus and human cytomegalovirus and identify pentameric symmetry as a unifying structural feature. We propose that the packaging accessory factor oligomerizes to concentrate positively charges in a central channel critical for packaging progression.

microbiology↗

bioRxiv2026

Programmed Double-stranded RNA Formation Enables Meiotic Stage Transitions

Cell fate transitions rely on extensive transcriptional activation mediated by transcription factors, yet the equally important processes that ensure the selective removal of pre-existing mRNAs remain elusive. We uncover widespread formation of double-stranded RNA (dsRNA) during early-to-middle meiosis using quantitative RNA structure analysis, and validate hundreds of natural antisense transcripts (NATs) through long-read sequencing. These NATs are induced by meiosis-specific transcription factors, including Ndt80p, and pair with sense mRNAs to drive cytoplasmic aggregation of the resulting dsRNAs. These dsRNA aggregates are subsequently transported to the vacuole for clearance, likely via autophagy. This pathway selectively eliminates mRNAs before metaphase I, including NDJ1, which encodes a meiotic cohesion protein that would otherwise block chromosome segregation. Our study reveals a physiological role for large-scale dsRNA formation, and highlights a simple but powerful model: the same transcription factors that activate mRNAs required for a specific developmental stage also program the synthesis of NATs that promote removal of transcripts from the preceding stage, thereby driving bidirectional transcriptome reprogramming.

cell biology↗

bioRxiv2026

RNA Structure Coordinates Translation Across the Meiotic Program

mRNA structure clearly modulates translation efficiency for individual transcripts, yet its role in coordinating translation across many transcripts remains poorly defined. Meiosis offers a compelling test case, involving large-scale gene expression changes coordinated across hundreds of mRNAs under conditions where transcription is constrained by chromosome condensation. We profiled mRNA structures across yeast meiosis, generating a high-resolution structurome encompassing ~70% of annotated mRNAs, including multi-time-point measurements for 2,084 transcripts. Transcripts upregulated during meiosis generally adopt flexible structures that enhance translation, whereas mRNAs with complex 5' UTR and coding-region structures show suppressed translation, indicating that RNA structure globally shapes meiotic translation. We further observed a high-low-high oscillation in cytoplasmic RNA helicase levels across meiosis. Together, these findings support a model in which RNA structure and helicase expression act jointly to program stage-specific translation of hundreds of mRNAs, with highly structured transcripts selectively translated at late stages. Consistent with this model, disrupting the structure of the abundant transcript CCW22, or altering Ded1p helicase levels, impaired meiotic progression in an RNA structure-dependent manner. Our study reveals that the concerted action of RNA structure and helicases coordinates cell-wide translation dynamics to meet stage-specific demands, especially critical when transcription is limited during meiotic divisions.

genomics↗

bioRxiv2026

A Hybrid Gag Lattice as a Structural Intermediate in HIV-1 Maturation

HIV-1 maturation converts the spherical immature Gag lattice into the conical mature capsid required for infectivity, yet the structural route linking these two lattice endpoints remains unclear. Using single-particle cryo-electron microscopy and cryo-electron tomography on reconstituted assemblies and virus-like particles (VLPs), we identify a distinct hybrid lattice in which the capsid protein (CA) N-terminal domain adopts a mature-like conformation, whereas the CA C-terminal domain-SP1 layer remains immature. This architecture is observed in native, enveloped VLPs produced in human cells, demonstrating formation under physiologically relevant conditions. The hybrid lattice engages three myo-inositol hexakisphosphate (IP6) molecules per CA-SP1 hexamer, underscoring IP6-dependent stabilization; accordingly, excess IP6 enriches the hybrid population. Tomographic mapping shows that hybrid regions coexist with the immature lattice within the same particle and are enriched near lattice discontinuities, consistent with edge-localized remodeling that can accommodate conformational rearrangements and partial disassembly and reassembly. Disrupting a hybrid-specific inter-hexamer contact preserves immature lattice assembly and particle release but abrogates infectivity, compromises core integrity, and prevents mature lattice formation, implicating the hybrid architecture as an on-pathway intermediate. Molecular dynamics simulations further support coordinated rearrangements that bias the system away from the immature configuration toward hybrid and then mature organizations. Together, these results support a maturation model in which localized displacive remodeling and partial disassembly/reassembly act in concert, with IP6 tuning the balance among lattice states. This study provides new insight into HIV-1 maturation and identifies the hybrid lattice as a potential therapeutic target.

biochemistry↗

bioRxiv2026

Pan-cancer silencer transcription atlas reveals leukemia silencer hijacking FOXP1 to enhance MYC oncogene

Numerous non-coding regulatory elements have been identified in the human genome. However, our understanding of silencers remains largely incomplete. Here, we proposed the concept of silencer transcriptional activity (STA) to investigated silencers from a new perspective using >20,000 transcriptomes. We demonstrated that lower STAs predicted better patient outcomes in most cancers, and a subset of STAs distinguished tumors from noncancerous tissues for most cancer types. Importantly, we discovered a silencer-linked RNA (slRNA) of a leukemia cell-specific silencer (LeSSi). Knocking down (KD) of the slRNA significantly decreased leukemia cell viability and migration and increased cellular apoptosis. Furthermore, the direct link and functional similarity between LeSSi and slRNA were established by CRISPR/Cas9 knocking out (KO) of LeSSi. Mechanistically, the slRNA hijacked FOXP1, which functions here both as a known transcription factor (TF), and more importantly, as a novel RNA binding protein (RBP), thereby preventing FOXP1 from binding to the promoter of proto-oncogene MYC and subsequently promoting MYC transcription. Collectively, our work provides insights into the functions of silencers in cancer and identified a slRNA hijacking FOXP1 to enhance MYC expression in leukemia cells.

genomics↗

bioRxiv2026

Pan-cancer RNA editing activity reveals complex editing functions and cancer immunotherapy biomarkers

Pervasive RNA editing in human diversifies the transcriptome and proteome. However, biological functions of most RNA editing sites remain to be uncovered. Here, we developed a computational framework (iPEAPR) to investigate editing functions in various pathways and regulatory elements, by using only summary information of millions of editing sites, in >5,000 cancer samples and in >2,500 normal samples, and large numbers of manually curated functional annotations. We observed heterogeneous editing activities across features. Surprisingly, enhancers were among features showing the highest editing activities. Editing of epithelial-mesenchymal transition was the most significantly associated with patient survival. Moreover, editing associations with cancer stemness, DNA repair deficiency, and tumor immune infiltrations uncovered known and potential regulators of tumor features. We constructed an editing-mediated regulatory network, which revealed new editing modulators, including experimentally validated TNRC6A. Lastly, we demonstrated that EIs can act as biomarkers of both anti-PD1 immunotherapy and other cancer drugs, such as MEK and BRAF inhibitors. Collectively, iPEAPR enabled depicting RNA editing-dependent abnormal functions in cancer and can be applied to more diseases.

bioinformatics↗

bioRxiv2025

Mast cell extracellular granules are bioactive condensates driven by heparin and polyamine

Biomolecular condensates are membraneless bodies that organize biochemical reactions typically within cells. However, the roles of condensates in extracellular space--where conditions differ substantially from intracellular space--remain poorly understood. Here, we report mast cell extracellular granules (MCEGs), a stable membraneless entity, are condensates assembled via electrostatic interactions between glycosaminoglycans and polyamines. Disrupting polyamine synthesis or trafficking blocks MCEG formation and compromises the storage of proteases and cytokines. Granules reconstituted with heparin and spermine are sufficient to enrich mediators such as CPA3 and TNF, maintaining an elevated pH and higher concentrations of calcium and zinc compared to the extracellular milieu. This unique environment enhances CPA3 enzymatic activity. Furthermore, the granules increase TNF binding and its bioactivity toward endothelial cells. Together, we reveal MCEGs as functionally active biomolecular condensates with distinct biochemical and immunological properties; MCEGs are formed through sugar-metabolite interactions, expanding the mechanisms of condensate assembly beyond classical protein-protein and protein-RNA interactions.

cell biology↗

bioRxiv2025

Fully computational design of PAM-relaxed Staphylococcus aureus Cas9 with expanded targeting capability

CRISPR-Cas9 nucleases have transformed genome engineering, yet their application is often constrained by protospacer-adjacent motif (PAM) requirements. Staphylococcus aureus Cas9 (SaCas9) is particularly attractive for in vivo applications due to its compact size; however, its NNGRRT PAM limits targetable genomic sites. Here, we report KRH, a SaCas9 variant designed entirely from the wild-type enzyme through a fully computational point-mutation design workflow, UniDesign, without additional experimental optimization. As expected, KRH efficiently recognizes an expanded NNNRRT PAM and exhibits substantially enhanced editing efficiency at non-canonical PAM sites, with improvements of up to 116-fold over the wild type. KRH achieves genome- and base-editing efficiencies comparable to, or exceeding, those of the well-known evolution-derived KKH variant. Computational modeling by UniDesign provides a mechanistic explanation for the PAM relaxation observed in both KRH and KKH, with structural and energetic analyses revealing that KRH relaxes PAM specificity by fine-tuning the balance between sequence-specific interactions with PAM bases and nonspecific contacts with the DNA backbone. Beyond its practical utility, KRH demonstrates that computational design can identify a minimal set of mutations sufficient to remodel the PAM interface while preserving high nuclease activity. This approach recapitulates--and in some cases surpasses--the performance of evolution-derived variants, offering a scalable strategy for high-throughput Cas9 engineering. Overall, these results establish KRH as a blueprint for rationally engineered, PAM-relaxed nucleases and underscores the power of computational design to accelerate next-generation genome editing.

bioengineering↗

bioRxiv2025

Structural Basis of TACO1-Mediated Efficient Mitochondrial Translation

Translation elongation is a universally conserved step in protein synthesis, relying on elongation factors that engage the ribosomal L7/L12 stalk to mediate aminoacyl-tRNA delivery, accommodation, and ribosomal translocation. Using in organello cryo-electron microscopy, we reveal how the mitochondrial translation accelerator TACO1 promotes efficient elongation on human mitoribosomes. TACO1 binds the mitoribosomal region typically bound by elongation factor Tu (mtEF-Tu), bridging the large and small subunits via contacts with 16S rRNA, bL12m, A-site tRNA, and uS12m. While active throughout elongation, TACO1 is especially critical when translating polyproline motifs. Its absence prolongs mtEF-Tu residence in A/T states, causes persistent mitoribosomal stalling and premature subunit dissociation. Structural analyses indicate that TACO1 competes with mtEF-Tu for mitoribosome binding, stabilizes A-site tRNA, and enhances peptidyl transfer through a mechanism distinct from EF-P and eIF5A. These findings suggest that bacterial TACO1 orthologs may serve analogous roles, highlighting an evolutionarily conserved strategy for maintaining elongation efficiency during challenging translation events.

biophysics↗

bioRxiv2025

Motility Functions as an Essential Defense in the Bacterial Persister Lifecycle

Bacterial persisters are phenotypically antibiotic-tolerant variants driving recurrent infections. While motility is a key trait for bacterial survival and virulence, its function throughout the persister lifecycle remains poorly understood. Here, we unveil a stage-specific trajectory for motility that underlies sophisticated survival strategies in rifampin-induced Escherichia coli persistence. During persister formation, intact motility functions as an immediate behavioral defense that decreases intracellular bactericidal antibiotic concentrations to ensure survival. This initial behavioral defense provides critical time for transcriptional reprogramming in persisters, which, by downregulating the major outer membrane porins (OmpC & OmpF) that facilitate antibiotic influx, supplants the preceding motility-based strategy. Subsequently, in structurally fortified persisters, motility becomes dispensable and gradually diminishes- a process independent of cell bioenergetic profile-yet re-emerges as essential for timely resuscitation upon antibiotic removal. Collectively, these findings establish the significance of motility in bacterial survival, redefine persister defense against antibiotics as a transition from behavioral to architectural strategies, and identify motility as a potential adjuvant therapeutic target. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=98 SRC="FIGDIR/small/694804v1_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@afdee5org.highwire.dtl.DTLVardef@52b4beorg.highwire.dtl.DTLVardef@1d865b6org.highwire.dtl.DTLVardef@1fd9cfb_HPS_FORMAT_FIGEXP M_FIG C_FIG The dynamic roles and underlying mechanisms of bacterial motility throughout the rifampin-induced persister lifecycle: motility initially serves as a behavioral defense mechanism essential for persister formation, then becoming dispensable in the persister state which implements a structural defense via membrane porin downregulation, and ultimately re-emerges as a critical requirement for timely resuscitation.

microbiology↗

bioRxiv2025

Synchronized Latency Reversal and Immune Clearance by a Multifunctional Fusion Protein Enables HIV-1 Reservoir Reduction

Strategies that concurrently reactivate latent reservoirs and enhance immune-mediated clearance hold significant promise for achieving an HIV cure. Here, we developed hyperIL-15xsCD4-Fc (15xsCD4-Fc), a fusion protein that integrates latency reactivation, targeted immune engagement, and effector-mediated killing into a single molecule. This agent not only potently reverses HIV-1 latency in CD4+ T cells from people living with HIV-1 (PLWH) through coordinated IL-15 receptor signaling and sCD4-mediated HIV-1 envelope (Env) engagement, but also enhances antigen-specific CD8+ T cell response in PBMCs derived from PLWH. Furthermore, 15xsCD4-Fc enables Env-specific elimination of reactivated latently infected cells by NK cells while preventing off-target cytotoxicity. In PBMCs from PLWH, 15xsCD4-Fc reduced replication-competent HIV-1 DNA by 93.8%. In antiretroviral treated HIV-1-infected humanized mice, the molecule demonstrated both safety and efficacy in diminishing viral reservoir in lymphoid organs. This spatiotemporally coupled approach to reservoir exposure and immune recognition establishes a clinically viable strategy for clearing the HIV-1 reservoir. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=195 SRC="FIGDIR/small/693434v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@8f8f0org.highwire.dtl.DTLVardef@10f63f7org.highwire.dtl.DTLVardef@16c1de2org.highwire.dtl.DTLVardef@11e1787_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Single-Cell Immune Profiling Reveals Non-classical Monocyte-Driven Immunodepression in Tuberculosis Treatment Non-Responders

Sputum culture conversion (SCC) at 2 months is a critical predictor of tuberculosis (TB) treatment outcomes, yet the underlying immune alterations driving treatment non-response remain unclear. We performed single-cell RNA sequencing on peripheral blood mononuclear cells from 8 TB patients, stratified by 2-month SCC status, to characterize cellular heterogeneity and intercellular communication. Treatment non-responders exhibited marked expansion and terminal differentiation of non-classical monocytes with elevated TB progression risk signatures. These cells showed reduced outgoing but enhanced incoming signaling, prominently via IL-16 and TRAIL pathways, potentially amplifying regulatory T cell (Treg)-mediated immunosuppression. CD4 Tregs in non-responders engaged in intensified GALECTIN signaling toward CD8 cytotoxic T cells and mature NK cells, contributing to effector cell exhaustion. NK cell subsets, particularly mature NK c2, displayed increased inhibitory receptor expression and terminal differentiation states. Our findings delineate a dysregulated "non-classical monocyte -Treg -cytotoxic lymphocyte" axis in TB treatment failure, highlighting candidate biomarkers and host-directed therapeutic targets.

bioinformatics↗

bioRxiv2025

ACE2 Phosphorylation Modulates Angiogenesis via the Activator Protein-1

BackgroundAngiogenesis plays a crucial role in organ development. However, aberrant blood vessel growth is involved in various diseases, including tumors and neovascular eye diseases. Angiotensin-converting enzyme 2 (ACE2) is a critical enzyme regulating the health of cardiovascular system, and its post-translational modifications (PTMs) are crucial to determine ACE2 expression level and activity. Here, we studied how the PTM of ACE2 in vascular endothelial cells (ECs) affect pathological retinal neovascularization and tumor angiogenesis. MethodsThe angiogenic capabilities of ECs were assessed by tube formation, sprouting assays, and 5-ethynyl-2-deoxyuridine (EdU) and filopodia staining. EC angiogenesis was examined by poteome profiler array and aortic ring assays in vitro and by the oxygen-induced retinopathy (OIR) and tumor angiogenesis models in vivo. High-throughput screening involving data from RNA-seq, ATAC-seq, and ChIP-seq were used to explore the epigenetic and transcriptional regulations of pro-angiogenic genes regualtged by ACE2 PTMs. ResultsACE2 Ser-680 dephosphorylation in connection with Lys-788 ubiquitination increased EC angiogenic phenotype, which were manifested by aberrant vascularization in mouse OIR and tumor models. ACE2 Ser-680 dephosphorylation led to the activation of activator protein-1 (AP-1), which transactivated multiple genes involved in angiogenesis. AP- 1 inhibition mitigated such angiogenesis in vivo. ConclusionOur findings show a novel PTM mechanism of ACE2 involved in pathological angiogeneis. Specifically, ACE2 Ser-680 dephosphorylation facilitated AP-1 transactivation of the downstream pro-angiogenic genes in ECs.

cell biology↗

bioRxiv2025

Late-Talking Children Talk More? A Machine Learning Approach to Speech Act Analysis in Early Childhood

Speech acts shape early language development and social cognition, yet little is known about how late-talking (LT) children use them to achieve communicative goals. We compared LT and typically developing (TD) preschoolers (1;09-6;00) across nine dyadic English corpora, using a Conditional Random Field model to annotate speech acts. We analyzed speech act distributions, hierarchical relations, and contingent responses to assess production and comprehension. TD children produced more declarative statements and wh-questions, whereas LT children produced more unclear word-like utterances and showed reduced comprehension ability (1;09-2;07). Speech acts classified LT and TD groups with 72.3% accuracy, improving to 76.6% with linguistic and demographic features. Classification was driven by co-occurring patterns of speech act frequencies. LT children showed delayed onset of speech acts but employed more speech acts after 3;09, focusing on speaker-centered goals, whereas TD children favored collaborative use of speech acts, revealing complex dynamics in the development of communicative skills.

pathology↗

bioRxiv2025

Theory of Mind and Discourse Production in Schizotypy: An fMRI Study

BackgroundSchizotypy (ST) reflects subclinical traits linked to schizophrenia spectrum disorders and associated cognitive and social impairments. Theory of Mind (ToM) and discourse production deficits are well-documented in schizophrenia (SZ), yet the neural basis of discourse-related ToM processes in ST remains unclear. This study investigated brain activation during narrative planning and production in individuals with schizotypal traits. MethodsThirty young adults (mean age = 18.8 years) completed standardized assessments, including the Schizotypal Personality Questionnaire-Brief Revised (SPQ-BR), adverse childhood experiences (ACEs), depression (PHQ-9), and dissociation (DES-B). Participants performed a discourse task in an fMRI scanner, describing nine-panel cartoons requiring inference of character intentions. Behavioral discourse metrics included total and inferred events. fMRI analyses examined activation during planning and production phases, with SPQ-BR positive, negative, and disorganized traits entered as regressors. ResultsSchizotypal traits correlated with multiple psychosocial risk factors, including elevated depression, ACEs, and dissociation (r = .48-.82, p < .01). During planning, canonical ToM/self-referential regions (vmPFC, precuneus, insula) were recruited. Positive traits correlated with increased activation in the right temporo-parietal junction, precuneus, and lingual gyrus, whereas disorganized traits were associated with reduced activation in the precuneus and lingual gyri. During production, networks spanning vmPFC, hippocampus, right TPJ, and basal ganglia were engaged. Negative traits correlated with increased motor/premotor activation, while disorganized traits correlated with reduced activation in lingual gyrus, SMA, and cerebellum. ConclusionsFindings demonstrate distinct neural correlates of schizotypal traits during discourse planning and production, supporting models of schizophrenia-spectrum risk emphasizing disrupted inference and integration processes.

neuroscience↗

bioRxiv2025

Integrative Systems Biology Identifies the AHR Signaling Axis as a Mechanistic Basis for Xiaobi Plaster in Lumbar Disc Degeneration

BackgroundLumbar disc degeneration (LDD) is characterized by chronic inflammation, oxidative stress, and extracellular matrix (ECM) breakdown, yet current therapies provide only symptomatic relief. Xiaobi Plaster (XBG), a traditional Chinese transdermal herbal preparation, has shown clinical benefit, but its molecular mechanisms remain unclear. MethodsWe applied an integrative systems biology framework combining network pharmacology, machine learning, and transcriptomic validation to identify candidate targets of Xiaobi Plaster in LDD. Molecular docking, dynamics simulations, and immune infiltration analysis further elucidated the underlying mechanisms, which were experimentally validated in a rat puncture-induced degeneration model. ResultsThree key targets--AHR, PTPN2, and SOAT1--were identified, with AHR emerging as the central regulator. Differential expression and enrichment analyses highlighted inflammatory and ECM-remodeling pathways, which overlapped with AHR-specific GSEA, underscoring its hub role in integrating immune and redox signaling with matrix turnover. Molecular docking and 100-ns molecular dynamics confirmed rutaecarpine (MOL002662) as the most stable AHR ligand. Transcriptomic data further showed AHR expression correlates with degeneration severity and mast cell infiltration. In vivo qPCR and Western blotting validated upregulation of AHR and supporting roles of PTPN2 and SOAT1 in degenerative discs. ConclusionThis study elucidates the multi-target and multi-pathway mechanisms of XBG in LDD, highlighting AHR as a pivotal therapeutic target modulated by rutaecarpine, with PTPN2 and SOAT1 serving as auxiliary regulators. These findings provide mechanistic insights into the transdermal application of XBG and support the therapeutic relevance of targeting the AHR signaling axis in LDD.

bioinformatics↗

bioRxiv2025

Dissecting the unique self-assembly landscape of the HIV-2 capsid protein

Human immunodeficiency virus type 2 (HIV-2) is a lentivirus closely related to HIV-1 but exhibits distinct molecular and clinical features that influence viral infectivity and efficacy of antiretroviral therapy. The HIV capsid is a critical structural component with multifaceted roles during infection and mediates some of the observed divergence between HIV-1 and HIV-2. Unlike HIV-1, study of the HIV-2 capsid is limited and standard protocols for the in vitro assembly of HIV-1 capsid protein (CA) lattice structures have not been successfully translated to the HIV-2 context. This work identifies effective approaches for the assembly of the HIV-2 CA lattice and leverages this to biochemically characterize HIV-2 CA assemblies and mutant phenotypes. Our findings elaborate on the sensitivity of HIV-2 CA to chemical conditions and reveal that it assembles into a more varied spectrum of particle morphologies compared to HIV-1. Utilizing these assemblies, we tested the hypothesis that HIV-1 and HIV-2 employ divergent mechanisms to stabilize CA oligomer forms and investigate the effects of non-conserved substitutions at the CA inter-protomer interfaces. This work advances our understanding of the key biochemical determinants of HIV-2 CA assembly that are distinct from HIV-1 and may contribute to their divergent virological properties.

biochemistry↗

bioRxiv2025

Inactivation of peptidoglycan remodeling promotes antibiotic susceptibility in vancomycin-resistant Enterococcus faecium

Vancomycin-resistant Enterococcus faecium (VREfm) is a leading cause of healthcare-associated infections globally and demands new approaches for treatment. Here we show that genetic and pharmacological inactivation of a highly conserved NlpC/P60 peptidoglycan hydrolase, secreted antigen A (SagA), enhanced vancomycin susceptibility of VREfm ex vivo and in vivo. Notably, genetic deletion of sagA impaired VREfm peptidoglycan remodeling, growth and increased the activity of vancomycin. We then identified first-in-class covalent NlpC/P60 peptidoglycan hydrolase inhibitors and demonstrated that pharmacological inactivation of SagA activity also impaired peptidoglycan remodeling and increased the efficacy of vancomycin across genetically distinct VREfm clinical isolates. Our study reveals peptidoglycan hydrolases are druggable targets whose inactivation improves the efficacy of vancomycin against VREfm.

microbiology↗