Co-option of a degenerated polysaccharide transporter for type VI protein secretion across diverse α-Proteobacteria

Co-option of a degenerate polysaccharide transporter replaces a core component of the type VI secretion system in α-Proteobacteria | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Co-option of a degenerate polysaccharide transporter replaces a core component of the type VI secretion system in α-Proteobacteria Li-Kang Sung, Jer-Sheng Lin, Yi-Chieh Wang, Chih-Feng Wu, Hao-Wen Hsu, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7993693/v2 This work is licensed under a CC BY 4.0 License Status: Posted Version 2 posted You are reading this latest preprint version Show more versions Abstract Type VI secretion system (T6SS) is a widespread bacterial nanomachine that mediates interbacterial antagonism and ecological competitiveness. Thirteen core type six secretion (Tss) proteins, including the outer membrane-anchoring protein TssJ, constitute the canonical T6SS. Yet, TssJ is absent from T6SSs in several lineages, raising the question of how their systems assemble and function without it. Using the TssJ-lacking T6SS of Agrobacterium C58 as a model, we describe a form of co-optive evolution in which a degenerated outer membrane polysaccharide transporter, TssX, functionally replaces TssJ. We delineated the assembly pathway of the agrobacterial T6SS and uncovered an assembly order in which TssX acts downstream of the membrane complex and prior to contractile sheath polymerization. Comparative genomic analyses of 32,414 complete proteobacterial genomes revealed that tssX defines a distinct T6SS subtype and is mutually exclusive with tssJ, indicating functional compensation through evolutionary replacement. Phylogenetic and synteny analyses revealed multiple independent losses of tssJ and subsequent co-option of tssX in α-proteobacterial lineages. Our work uncovers a novel T6SS outer membrane protein and shows that essential nanomachine elements can be replaced. The variation in both composition and assembly mechanisms exemplifies the remarkable flexibility in the evolution of protein complexes that are both structurally and functionally conserved. Biological sciences/Microbiology/Bacteria/Bacterial secretion Biological sciences/Evolution/Molecular evolution Full Text Additional Declarations The authors declare no competing interests. Supplementary Files TssXmanuscriptTableS1final.xlsx Supplementary Table 1 Mov.S1.mp4 Supplementary Mov. 1 Mov.S2.mp4 Supplementary Mov. 2 Mov.S3.mp4 Supplementary Mov. 3 Cite Share Download PDF Status: Posted Version 2 posted You are reading this latest preprint version Show more versions Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7993693","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":544360663,"identity":"4615725b-3410-4ca5-b34b-d96eb8251b2e","order_by":0,"name":"Li-Kang Sung","email":"","orcid":"","institution":"Institute of Plant and Microbial Biology, Academia Sinica","correspondingAuthor":false,"prefix":"","firstName":"Li-Kang","middleName":"","lastName":"Sung","suffix":""},{"id":544360664,"identity":"a5f5ff03-158d-43bc-b639-7a70c72feaa3","order_by":1,"name":"Jer-Sheng Lin","email":"","orcid":"","institution":"Academia 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α-Proteobacteria\u003c/strong\u003e\u003c/p\u003e","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research 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Thirteen core type six secretion (Tss) proteins, including the outer membrane-anchoring protein TssJ, constitute the canonical T6SS. Yet, TssJ is absent from T6SSs in several lineages, raising the question of how their systems assemble and function without it. Using the TssJ-lacking T6SS of Agrobacterium C58 as a model, we describe a form of co-optive evolution in which a degenerated outer membrane polysaccharide transporter, TssX, functionally replaces TssJ. We delineated the assembly pathway of the agrobacterial T6SS and uncovered an assembly order in which TssX acts downstream of the membrane complex and prior to contractile sheath polymerization. Comparative genomic analyses of 32,414 complete proteobacterial genomes revealed that tssX defines a distinct T6SS subtype and is mutually exclusive with tssJ, indicating functional compensation through evolutionary replacement. Phylogenetic and synteny analyses revealed multiple independent losses of tssJ and subsequent co-option of tssX in α-proteobacterial lineages. Our work uncovers a novel T6SS outer membrane protein and shows that essential nanomachine elements can be replaced. 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