mBio最新文献

{"title":"Redox control and mechanisms of transmembrane signaling in CSS domain c-di-GMP phosphodiesterases that control biofilm formation in <i>Escherichia coli</i>.","authors":"Katharina Pressler, Martin Lorkowski, Regine Hengge","doi":"10.1128/mbio.01325-26","DOIUrl":"10.1128/mbio.01325-26","url":null,"abstract":"<p><p>Bis-(3',5')-cyclic dimeric guanosine monophosphate (c-di-GMP) control of biofilm formation in <i>Escherichia coli</i> K-12 is balanced by multiple diguanylate cyclases (DGCs) and c-di-GMP-specific phosphodiesterases (PDEs). Five of the 13 PDEs feature a periplasmic CSS domain with two conserved cysteines, flanked by transmembrane (TM) regions, and an enzymatically active cytoplasmic EAL domain. One of these (PdeC) was previously shown to be redox-regulated by DsbA/DsbB-mediated disulfide bond (DSB) formation in the cysteine serine serine (CSS) domain. Comparing all five CSS domain PDEs, we found them to fall into two groups with similar biochemical features, resulting in different consequences for PDE activity. PdeB, PdeC, and PdeG are more active when lacking the periplasmic DSB, resulting in diminished biofilm formation, while PdeN and PdeD are active in their oxidized forms. Using PdeB and PdeN as prototypes for the two groups, not only the periplasmic DSB but also differently charged amino acid motifs close to the transmembrane (TM) domains and a putative additional DSB in PdeN were identified as important for transmembrane signaling. All these elements, including its stable structural DSB in the periplasm, which can form independently of DsbA, maintain PdeN in a rigid, proteolysis-resistant active conformation. By contrast, for PdeB, the more rigid DSB-containing conformation is inactive, with mutations in the stabilizing elements leading to a structurally less constrained, more active enzyme. Notably, low PdeN levels are post-transcriptionally upregulated at acidic pH, resulting in less biofilm formation. Overall, the five CSS domain PDEs enable <i>E. coli</i> to adapt to diverse environmental niches.IMPORTANCESensing environmental cues and transmembrane signal transduction via membrane-embedded proteins is a process of key importance in all living cells. To investigate the molecular mechanisms involved, we performed a systematic functional comparison of the five CSS domain phosphodiesterases of <i>Escherichia coli</i>, which degrade the bacterial second messenger c-di-GMP in response to redox and other signals. With a sensory domain in the periplasm linked to a cytoplasmic enzymatic domain, these proteins represent minimal devices for transmembrane signaling. We demonstrate that these signal-transducing enzymes fall into two functional classes with a similar periplasmic redox biochemistry resulting in opposite states of cytoplasmic enzymatic activity. Several characteristic sequence elements convey redox and structural information in the periplasmic and transmembrane protein segments to their ability to dimerize into an enzymatically active form in the cytoplasm. Comparing the five enzymes also shows that evolution has played with these elements to facilitate adaptation to various environmental niches.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0132526"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148865595","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"KSHV promotes leukocyte adhesion and transendothelial migration via induction of VCAM1.","authors":"Zhigang Zhang, Kurtis M Host, Jason Wong, Penny Anders, Blossom Damania","doi":"10.1128/mbio.01499-26","DOIUrl":"10.1128/mbio.01499-26","url":null,"abstract":"<p><p>Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus are gammaherpesviruses associated with multiple human cancers. Kaposi's sarcoma (KS) is a human malignancy associated with KSHV infection. KS lesions are characterized by proliferating endothelial-derived spindle cells, leaky blood vessels, and inflammatory infiltrating leukocytes. The inflammatory leukocytes secrete cytokines and growth factors that may contribute to the survival and proliferation of tumor cells. Unless exposed to inflammatory cytokines, resting endothelial cells usually do not express adhesion molecules. Here, we identify a novel mechanistic link demonstrating that KSHV-infected latent endothelial cells express vascular cell adhesion molecule (VCAM1) and induce leukocyte adhesion and transendothelial migration (TEM) in a VCAM1-dependent manner. Inhibition of VCAM1 function using siRNA knockdown or antibody blockade impairs leukocyte adhesion and TEM, suggesting that cell adhesion and TEM are mediated by VCAM1. VCAM1 expression in KSHV-infected endothelial cells involves the non-canonical NF-кB (NF-κB2) pathway because depletion of NIK, IKKα, RelB, or NF-κB p52 via siRNA results in a decrease of VCAM1 levels, as well as a corresponding impairment of cell adhesion and TEM. Thus, KSHV-infected endothelial cells likely promote the infiltration of immune cells by modulating the expression of adhesion factors like VCAM1 on their surface. Moreover, we report that KSHV vFLIP, a latency protein encoded by the virus, directly induces VCAM1 expression and promotes leukocyte adhesion and transendothelial migration. We further demonstrate that endothelial cells infected with a vFLIP-deleted virus had reduced VCAM1 expression and reduced leukocyte adhesion and transendothelial migration.IMPORTANCEKaposi's sarcoma-associated herpesvirus (KSHV) is associated with the development of Kaposi's sarcoma (KS). KS tumors are characterized by proliferating endothelial-derived spindle cells and infiltrating leukocytes, which secrete cytokines and growth factors that may contribute to the proliferation of tumor cells. Unless exposed to inflammatory cytokines, resting endothelial cells usually do not express adhesion molecules. Here, we demonstrate that KSHV-infected latent endothelial cells induce the expression of vascular cell adhesion molecule, which promotes leukocyte adhesion and transendothelial migration.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0149926"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148906735","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The cellular protein TIAR mediates rapid initiation of West Nile virus genome RNA synthesis.","authors":"Jessica L Siemer, Hsuan Liu, Jin Zhang, Dan Cui, Jeenah Jung, Philip J Santangelo, Markus W Germann, Margo A Brinton","doi":"10.1128/mbio.01947-26","DOIUrl":"10.1128/mbio.01947-26","url":null,"abstract":"<p><p>During the intracellular replication cycle of West Nile virus (WNV), genome RNA synthesis is initially inefficient but increases exponentially as viral replication complexes are sequestered in invaginations in the endoplasmic reticulum. In this study, we investigated the functional role of the cellular protein TIAR (T-cell intracellular antigen-related protein) in the transcription of WNV genome RNA. Close colocalization of cytoplasmic TIAR with viral double-stranded RNA was detected by a proximity ligation assay in WNV-infected cells. TIAR binds specifically to the WNV 3'(-) SL but not to the complementary WNV 5'(+) SL in <i>in vitro</i> RNA binding assays. Only the 3' end of the WNV minus-strand RNA was enriched by immunoprecipitation of infected cell lysates with anti-TIAR antibody. Stable overexpression of TIAR in clonal A549 cells increased the ratio of intracellular viral plus-strand to minus-strand RNA in a dose-dependent manner. TIAR contains three RNA recognition motifs (RRMs). Biophysical data indicated that only RRM2 directly contacts RNA and that up to three TIAR molecules can bind cooperatively to the WNV 3'(-) SL RNA. These data provide additional evidence that TIAR functions as a proviral host factor facilitating exponential amplification of WNV genome production in infected cells.IMPORTANCEWest Nile virus (WNV) is a mosquito-borne orthoflavivirus associated with increasing global human disease incidence. The molecular mechanisms underlying viral replication are not fully understood. In early stages of infection, viral genome transcription is inefficient; however, in late stages, viral genome transcription increases exponentially. T-cell intracellular antigen-related (TIAR) protein is a cellular protein that has been shown to interact with the 3' end of the WNV negative-sense antigenomic RNA. We obtained data showing colocalization of cellular TIAR with viral replication complexes in infected cells and an increased ratio of intracellular genomic to antigenomic viral RNA in TIAR-overexpressing cells, and confirmed preferential binding of TIAR to the 3' end of the WNV antigenome both <i>in vitro</i> and in infected cell extracts. We also demonstrated that multiple TIAR proteins can bind cooperatively to the WNV 3'(-) stem-loop RNA. These data provide supporting evidence for a model of TIAR-mediated rapid initiation of nascent genome RNA synthesis in infected cells.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0194726"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148906790","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Monitoring kinetic changes and restriction of influenza A virus RNA species during infection using a Flu-Stranded CRISPR platform.","authors":"Anh-Tu Tran, Wen-Fang Tang, Chung-Fan Hsieh, Ya-Fang Chiu, Chung-Guei Huang, Kuo-Ming Lee, Rei-Lin Kuo, Kuei-Ching Hsiung, Cheng-Hsun Chiu, Jim-Tong Horng","doi":"10.1128/mbio.01789-26","DOIUrl":"10.1128/mbio.01789-26","url":null,"abstract":"&lt;p&gt;&lt;p&gt;Influenza A virus (IAV) generates three closely related RNA species: viral RNA (vRNA), complementary RNA (cRNA), and messenger RNA (mRNA), whose strand-specific quantification remains limited by sensitivity and quantitative dynamic range, particularly at low RNA abundance. Here, we developed Flu-Stranded CRISPR-Cas12a, a strand-specific detection platform integrating tagged reverse transcription, segment-specific PCR, and Cas12a collateral cleavage to support quantitative analysis of all three RNA species across a broad dynamic range. The assay enables reliable detection down to 10&lt;sup&gt;2&lt;/sup&gt; copies per reaction, extending the lower quantitative boundary relative to both SYBR Green and TaqMan reverse transcription quantitative PCR (RT-qPCR) under matched conditions. Validated in infected cell lines, murine lung tissues, and clinical nasopharyngeal specimens, the platform enabled subtype-discriminating, strand-resolved detection, including samples near or below the quantitative range of SYBR Green RT-qPCR. Using finely resolved infection time-course analyses in NP and NA segments, we identified a reproducible early vRNA decline within the early post-infection phase. This decline was partially attenuated in RIG-I knockout A549 cells, while subsequent vRNA accumulation was enhanced, consistent with a modulatory rather than essential role for RIG-I in early viral RNA dynamics. Subcellular fractionation localized this decline to cytoplasmic incoming genomes. In contrast, importazole-mediated inhibition of nuclear import abolished vRNA recovery without affecting the early decline, indicating that nuclear entry functionally separates early genome reduction from subsequent productive replication. These findings establish Flu-Stranded CRISPR-Cas12a as a strand-resolved framework for monitoring IAV RNA dynamics and reveal an early window of genome vulnerability during cytoplasmic transit that shapes infection outcome.&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Importance: &lt;/strong&gt;The early fate of incoming influenza virus genomes remains unclear, limiting our understanding of how infection is established or aborted in host cells. We developed Flu-Stranded CRISPR-Cas12a, a strand-specific platform for sensitive and quantitative analysis of influenza viral RNA (vRNA), complementary RNA (cRNA), and messenger RNA (mRNA) across experimental and clinical samples. Using high-resolution time-course analysis, we identified a reproducible early decline in vRNA during the post-entry phase. Our data suggest that this early genome loss arises from multiple processes, with RIG-I acting as a modulatory factor rather than a primary driver. Subcellular fractionation localized this effect to cytoplasmic incoming genomes, whereas importin-β-mediated nuclear entry was required for subsequent vRNA recovery. These findings support a model of an early cytoplasmic phase of genome attrition that is distinct from replication and provide a framework for understanding early influenza RNA kinetics and for","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0178926"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148963515","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The <i>Chlamydia trachomatis</i> effector CT226 directly engages LRRFIP1 to recruit a multiprotein complex that limits inclusion size and progeny production.","authors":"Cherilyn A Elwell, Amy Diallo, Barbara S Sixt, Jessica Sherry, Khavong Pha, Raphael H Valdivia, Oren Rosenberg, Kliment Verba, Joanne Engel","doi":"10.1128/mbio.01953-26","DOIUrl":"10.1128/mbio.01953-26","url":null,"abstract":"<p><p><i>Chlamydia</i> species secrete a unique class of effector proteins termed inclusion membrane proteins (Incs) that are translocated into the inclusion membrane and reprogram host cell processes to support intracellular growth. Using affinity purification-mass spectrometry, we previously identified an interaction between the early expressed Inc CT226 and a human multiprotein complex comprising leucine-rich repeat Flightless-I-interacting protein 1 (LRRFIP1), LRRFIP2, Flightless-I (FLII), and tropomodulin 3 (TMOD3). Here, we define the molecular basis and functional consequences of this interaction during <i>Chlamydia trachomatis</i> infection. We show that CT226 is both necessary and sufficient to recruit the LRRFIP1/2:FLII:TMOD3 (LFT) complex to the inclusion membrane. <i>In vitro</i> reconstitution with purified proteins demonstrates that the predicted coiled-coil region of CT226 directly binds the N-terminal helix of LRRFIP1. Using <i>in silico</i> structural modeling with AlphaFold-Multimer coupled with mutational validation, we identify key residues that mediate an electrostatic CT226:LRRFIP1 interaction required for LFT recruitment to the inclusion. Unexpectedly, genetic disruption of CT226 or its interaction with the LFT complex results in modest increases in inclusion size and recovery of infectious progeny late in infection, demonstrating that the CT226:LFT interaction modulates <i>C. trachomatis</i> developmental progression. Together, these findings establish the molecular basis of CT226-mediated recruitment of the LFT complex to the inclusion membrane and uncover a previously unrecognized role for this effector-host interaction at late timepoints of the <i>C. trachomatis</i> intracellular developmental cycle.IMPORTANCE<i>Chlamydia trachomatis</i> is a major cause of sexually transmitted infections and blindness worldwide. This bacterium survives by replicating inside host cells within a membrane-bound compartment, where it deploys specialized proteins to manipulate host processes. These proteins, known as inclusion membrane proteins (Incs), help organize the host-pathogen interface, but the molecular mechanisms by which many Incs engage host factors remain poorly understood. Here, we show that the Inc protein CT226 recruits a host multiprotein complex to the bacterial compartment by directly binding one of its components. Using structural modeling and targeted mutations, we define the molecular interface required for this interaction. Disruption of CT226 or its ability to engage this host complex results in modest increases in inclusion size and production of infectious progeny, suggesting a role for the CT226:host interaction in regulating <i>Chlamydia</i> intracellular development. These findings reveal how a bacterial effector can engage a host protein complex to influence the outcome of infection.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0195326"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148990506","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Experimental versus clinical studies capture distinct types of relapse in tuberculosis.","authors":"Christian T Michael, Maral Budak, Philana Ling Lin, Denise Kirschner","doi":"10.1128/mbio.01977-26","DOIUrl":"10.1128/mbio.01977-26","url":null,"abstract":"<p><p>Tuberculosis (TB) remains a global health concern, as <i>Mycobacterium tuberculosis</i> (Mtb) currently infects a quarter of the world's population. Though many TB patients cure infection with short-course treatment, regimens shorter than 4 months risk post-treatment relapse. This is facilitated by the presence of granulomas, spatially heterogeneous immune structures that form during infection that harbor a caseated core that sequesters Mtb in a non-replicating state and pharmacokinetically impedes antibiotics. Typically, clinical relapse is described as TB recurrence <2 years after a misdiagnosis of cure upon treatment completion (MDxC). Comparatively, many experimental models cannot (or do not) screen for cure and examine relapse ~2 months after treatment completion. Understanding how relapse occurs, and whether it is impacted by experimental versus clinical study designs, may improve treatment. Here, we examine two hypothesized mechanisms of relapse: (i) reservoir, where treatment kills all Mtb except those in a non-replicating, diagnostically inaccessible host reservoir; and (ii) threshold, where replicating Mtb remain alive below detectable levels within a test-accessible reservoir. We use our computational model capturing whole-host Mtb infection dynamics, HostSim. Our simulations uncover rates of reported relapse correlating with how studies are performed. Experimentally, relapse is likely driven by incomplete sterilization of replicating bacteria, whereas clinical relapse is most likely driven by gradual expansion of non-replicating Mtb from caseum into granuloma cellular areas. This suggests that TB patients who relapse may best be re-treated with regimens including caseum-penetrating antibiotics, such as rifamycin-class antibiotics, and that experimental animal relapse models better translate to clinical when \"cure\" is established at treatment completion.IMPORTANCEIncomplete treatment of TB leads to risks relapse, which may occur years later. The threat of relapse is the main reason TB treatment takes 4-9 months. Predictors of relapse are not well-defined given variability in technical definitions of relapse and nuances of study designs. Here, we use biologically based computation to simulate both clinical and experimental TB relapse studies, employing multiple diagnostic tests and relapse definitions. We find that two hypothesized types of relapse, each possibly requiring different re-treatment strategies, are simultaneously at play. Simulations suggest that relapse after a \"cure\" diagnosis (most often clinical studies) is caused by reactivation of non-replicating bacteria hidden from treatment within necrotic granuloma tissue or other sites, whereas experimental systems that cannot or do not test for cure post-treatment are likely to report relapse caused by incomplete sterilization of replicating Mtb. Predictions also depend on the currently unclear relationship between bacterial burden and clinical symptoms.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0197726"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148990552","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Altered physiology of drug-resistant <i>Mycobacterium tuberculosis</i> restricts adaptation to host-relevant conditions and produces condition-dependent collateral vulnerabilities.","authors":"William J Jowsey, Kate R Harding, Caitlin E Cunniffe, Chen-Yi Cheung, XinYue Wang, Caitlan J Smart, Hannah R Klaus, Amanda L Peterson, Brunda Nijagal, Mark B Hampton, Gregory M Cook, Matthew B McNeil","doi":"10.1128/mbio.00244-26","DOIUrl":"10.1128/mbio.00244-26","url":null,"abstract":"<p><p>Antibiotic resistance mutations often disrupt essential cellular processes; however, the physiological consequences of these mutations under host-relevant conditions remain poorly understood in many bacterial pathogens. Here, we have used a combination of phenotypic growth assays and metabolomics to investigate how drug resistance affects the metabolism of <i>Mycobacterium tuberculosis</i> and its ability to adapt to different host-relevant conditions. This work shows that (i) the fitness costs of antibiotic resistance in <i>M. tuberculosis</i> are highly variable and can be exacerbated or alleviated depending on the media carbon source, (ii) genetically unrelated resistance mutations have overlapping phenotypes and metabolic dysregulation, (iii) the growth of bedaquiline- and rifampicin-resistant mutants was impaired on host-relevant carbon sources, and (iv) resistance mutants have condition-specific collateral antibiotic susceptibilities, with rifampicin-resistant mutants being hypersusceptible to ATP synthase inhibitors when grown under conditions that require propionate metabolism. Collectively, this study highlights how drug resistance impacts bacterial physiology and places metabolic constraints on the ability of <i>M. tuberculosis</i> to adapt to changes in host-relevant conditions. This work has major implications for our understanding of how resistance evolves within the host context and reinforces the importance of how physiological variation in <i>M. tuberculosis</i> might affect the outcomes of new treatment regimens.</p><p><strong>Importance: </strong>Mutations that provide antibiotic resistance in bacterial pathogens often disrupt key cellular processes. The effects of these disruptions on cellular physiology are poorly understood, especially within the context of a host environment. In this article, we investigated how antibiotic resistance mutations affect the ability of <i>Mycobacterium tuberculosis</i>, the causative agent of tuberculosis, to grow under host-relevant conditions. Using techniques that characterize growth and metabolism, we demonstrate that antibiotic-resistant <i>M. tuberculosis</i> can have both increased and decreased growth depending on the media carbon source, highlighting how interactions between drug resistance traits and the environment affect the ability of <i>M. tuberculosis</i> to adapt to changing conditions. We also show that unrelated antibiotic-resistant strains share similar metabolic defects, which can be exploited to improve antimicrobial susceptibility. Overall, this work demonstrates that drug resistance reshapes the physiology of <i>M. tuberculosis</i> in growth-dependent ways, having major implications for our understanding of how drug resistance evolves within the host context and for the design of new treatment strategies against antibiotic-resistant <i>M. tuberculosis</i>.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0024426"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149039612","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A novel adenovirus 19K/IX protein enhances infection by attenuating proteasomal degradation of tyrosine-ubiquitinated capsid protein IX.","authors":"Erik Schubert, Vasileios Rafail Evangelopoulos, Mårten Larsson, Tanel Punga","doi":"10.1128/mbio.01683-26","DOIUrl":"https://doi.org/10.1128/mbio.01683-26","url":null,"abstract":"<p><p>Human adenoviruses (HAdVs) are double-stranded DNA viruses that cause a wide range of diseases, including respiratory, ocular, and gastrointestinal infections. HAdVs rely extensively on alternative splicing to expand their coding capacity and regulate gene expression during infection. Using replication-competent epitope-tagged HAdV-C5 viruses, we investigated the function of a newly identified alternatively spliced viral transcript, 19K/IX, derived from the <i>E1B19K</i> and <i>pIX</i> genes. We show that the 19K/IX protein binds to and stabilizes the minor capsid protein IX (pIX), thereby attenuating its proteasomal degradation and enhancing the production of infectious viral progeny. Mechanistically, we demonstrate that pIX degradation is mediated by non-canonical ubiquitination of the conserved tyrosine residues Y14 and Y49, which confer pIX subnuclear localization and partially mediate its interaction with the PSMC3 subunit of the 26S proteasome. Collectively, our study identifies 19K/IX as a novel regulator of HAdV-C5 infection and suggests that non-canonical tyrosine ubiquitination contributes to the regulation of capsid protein IX stability.</p><p><strong>Importance: </strong>Human adenoviruses (HAdVs) are clinically important pathogens and widely used as therapeutic tools. Because of their compact genomes, HAdVs use their genetic information very efficiently through alternative pre-mRNA processing. Here, we describe and characterize 19K/IX, a novel fusion protein generated by alternative splicing of two viral genes during infection. We demonstrate that 19K/IX stabilizes capsid protein IX by preventing its degradation, revealing a previously unrecognized strategy by which HAdVs ensure efficient production of new virus particles. These findings deepen our understanding of HAdV-C5 gene regulation and protein homeostasis and highlight the functional importance of non-canonical HAdV transcripts.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0168326"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149182874","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"HIV-1 hijacks HGF/c-MET signaling to promote viral entry and replication in primary CD4⁺ T cells.","authors":"Shraddha Tripathi, Madeleine M Allen, Li Wu","doi":"10.1128/mbio.02090-26","DOIUrl":"10.1128/mbio.02090-26","url":null,"abstract":"<p><p>The mesenchymal-epithelial transition factor (c-MET) is a receptor tyrosine kinase best known for mediating hepatocyte growth factor (HGF) signaling in cancer, but its role in HIV-1 infection has remained unclear. Here, we identify c-MET as a host factor that promotes HIV-1 replication in CD4⁺ T cells by facilitating viral entry. HIV-1 infection upregulates both c-MET transcript abundance and receptor phosphorylation in primary CD4⁺ T cells. Genetic knockout, knockdown, or pharmacological inhibition of c-MET in CD4⁺ T cells markedly reduced HIV-1 Gag protein expression, viral production, and infectivity. Integrative analyses using RNA sequencing, flow cytometry, and HIV-1 infection and entry assays revealed that c-MET disruption decreased cell-surface expression of the HIV-1 receptor CD4 and the coreceptor CXCR4, thereby limiting viral entry and subsequent infection. In addition, c-MET disruption attenuated activation of NF-κB, STAT1/3, and MAPK signaling pathways, which are critical regulators of HIV-1 gene expression. Together, these findings demonstrate that HIV-1 exploits HGF/c-MET signaling to coordinate entry receptor availability with intracellular signaling programs that support viral replication. Our study identifies a previously unrecognized host pathway required for efficient HIV-1 infection of primary CD4⁺ T cells and highlights HGF/c-MET signaling as a potential target for host-directed anti-HIV-1 therapeutic strategies.IMPORTANCEThe hepatocyte growth factor (HGF)/mesenchymal-epithelial transition factor (c-MET) signaling pathway is a well-established regulator of cell growth, survival, and migration and has been extensively studied as a therapeutic target in cancer. However, its role in HIV-1 infection has remained unknown. In this study, we found that c-MET enhances HIV-1 infection of primary CD4⁺ T cells by facilitating viral entry. We further demonstrate that HIV-1 replication enhances HGF-dependent c-MET phosphorylation, which is required for downstream signaling. Genetic ablation or pharmacologic inhibition of c-MET reduces HIV-1 entry by decreasing expression of viral entry receptors. Moreover, c-MET inhibition or knockout disrupts activation of cellular signaling pathways, including NF-κB, JAK/STAT, and p38 MAPK. These findings reveal a previously unrecognized role for HGF/c-MET signaling in HIV-1 pathogenesis and suggest that targeting c-MET may represent a novel host-directed strategy for HIV-1 intervention.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0209026"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149182884","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"HMGB1-targeted anti-inflammatory rBox A gene therapy for viral airway infections and systemic inflammatory disease models.","authors":"Kari Ann Shirey, Daniel Prantner, Lynda Coughlan, Chanhee Park, Lin Zou, Wei Chao, John Joseph, Jorge C G Blanco, Stefanie N Vogel","doi":"10.1128/mbio.01819-26","DOIUrl":"10.1128/mbio.01819-26","url":null,"abstract":"<p><p>High mobility group box 1 (HMGB1) is a host-derived proinflammatory molecule released during cell injury. Elevated extracellular HMGB1 levels have been associated with disease severity. HMGB1 contributes to host influenza responses by stimulating TLR4/MD-2 signaling, amplifying lung inflammation, and tissue damage via cytokine production and leukocyte infiltration. HMGB1 also plays a central role in other inflammatory diseases, including sepsis and liver toxicity, making it both a biomarker of disease severity and a potential therapeutic target. We previously reported development of a non-replicating adenovirus (AdV) vector that carries an inflammation-inducible cassette that remains dormant in the absence of inflammation but produces recombinant HMGB1 Box A, a competitive antagonist of HMGB1, in response to inflammatory stimuli. Therapeutic treatment with AdV.C3-Tat/HIV-Box A mitigated lung and systemic inflammation in response to influenza in mice and cotton rats. Herein, we have optimized vector delivery and expanded its use in multiple models of infectious and non-infectious HMGB1-induced inflammation.</p><p><strong>Importance: </strong>Despite promising preclinical outcomes, clinical translation of high mobility group box 1 (HMGB1) or TLR4/MD-2 antagonists for inflammatory diseases has been limited by challenges of drug synthesis, dosing, delivery, and off-target effects. Recombinant Box A (rBox A) is a competitive inhibitor of HMGB1 that binds to TLR4 and disrupts HMGB1-mediated TLR4/MD-2 signaling. rBox A exhibits broad anti-inflammatory activity across multiple diseases (e.g., endotoxemia, sepsis, neuroimmune disorders, ischemia/reperfusion); however, its therapeutic utility is constrained by the need for high, repeated dosing. We have engineered a non-replicating adenoviral (AdV) vector (AdV.C3-Tat/HIV-Box A) that produces recombinant (r)Box A when \"sensed\" by an inflammation-responsive complement C3 (C3) promoter. Therapeutic administration of AdV.C3-Tat/HIV-Box A resulted in significant protection and reduced inflammation in multiple HMGB1-mediated disease models, advancing a first-in-class, broadly applicable, host-directed gene therapy that selectively disrupts HMGB1-TLR4 signaling and has potential as both a broad-spectrum therapeutic for HMGB1-driven inflammatory diseases and a rapid-response countermeasure against emerging respiratory pathogens.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0181926"},"PeriodicalIF":5.4,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148808801","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}