Connexin Restrains EMT by Dual-Domain Mechanisms to Preserve Epithelial Identity

Connexin Restrains EMT by Dual-Domain Mechanisms to Preserve Epithelial Identity | 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 Article Connexin Restrains EMT by Dual-Domain Mechanisms to Preserve Epithelial Identity JEAN JIANG, Bo Ma, Zhaodan Ding, Guangyan Wang, Sumin Gu This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7501684/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Epithelial–mesenchymal transition (EMT) contributes to fibrotic disease in multiple organs and underlies post-surgical complications such as posterior capsule opacification (PCO), the leading cause of vision loss following cataract surgery. The molecular mechanisms that preserve epithelial integrity and restrain EMT remain poorly defined. Connexins are conventionally regarded as gap junction proteins mediating intercellular communication, but here we identify connexin 50 (Cx50) as a previously unrecognized key regulator of EMT. Using in vitro, ex vivo, and in vivo chick and mouse models, we show that transforming growth factor-β2 (TGF-β2) downregulates Cx50 and induces EMT markers including α-smooth muscle actin and fibronectin, changes reversed by Cx50 overexpression. Mechanistically, the extracellular E2 domain of Cx50 mediates adhesive interactions that limit epithelial migration and EMT, while the C-terminal domain stabilizes β-catenin–E-cadherin complexes and prevents TGF-β2–driven E-cadherin loss and β-catenin nuclear translocation. Dominant-negative E2 mutants abolish protection, and Cx50 knockout mice exhibit accelerated EMT after extracapsular lens extraction. These findings redefine connexins beyond their classical role as gap junction proteins, establishing Cx50 as a dual-domain regulator of EMT and epithelial plasticity with implications for fibrotic responses in ocular and other epithelial tissues. Biological sciences/Cell biology/Mechanisms of disease Health sciences/Diseases/Trauma Full Text Additional Declarations (Not answered) Supplementary Files OriginalDatafiles.pdf Original Data Files Supplementaldata.pdf Supplemental figures Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version 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-7501684","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":551234383,"identity":"06ec2c19-5ad3-4135-b24d-6b42fb4b8bcb","order_by":0,"name":"JEAN JIANG","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA0UlEQVRIiWNgGAWjYBACxmYQyWPDwCABEzpAnJY0Bh6itUDBYRK0MLczP3v4ReZ84n7p9ocff9QwyPHdSCDkMDZzYxme24k9MmeMpXmOMRhLEtbCYCYtAdIikcPGzMDGkLiBsBb2b0At54Ba0p8x/vjHUE+EFh4zyQ88B4BaEswYeNsYEgyI0FImzcCTbNxzI8dYmrdPwnDmmQf4tRj2H98m+bPHTrZ9RjowxL7ZyPMdJ2CLYQMwoHl74HwJ3EphQB7kuB8/CCscBaNgFIyCEQwAq8xBLLmsFVcAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0002-2185-5716","institution":"UNIVERSITY OF TEXAS HEALTH SCIENCE CENTER AT SAN ANTONIO","correspondingAuthor":true,"prefix":"","firstName":"JEAN","middleName":"","lastName":"JIANG","suffix":""},{"id":551234384,"identity":"50922c51-0079-4260-a906-ad3cdde1aa2d","order_by":1,"name":"Bo Ma","email":"","orcid":"","institution":"UNIVERSITY OF TEXAS HEALTH SCIENCE CENTER AT SAN ANTONIO","correspondingAuthor":false,"prefix":"","firstName":"Bo","middleName":"","lastName":"Ma","suffix":""},{"id":551234385,"identity":"421e326b-1757-4c91-9037-79691cf59653","order_by":2,"name":"Zhaodan Ding","email":"","orcid":"","institution":"University of Texas Health Science Center at San Antonio","correspondingAuthor":false,"prefix":"","firstName":"Zhaodan","middleName":"","lastName":"Ding","suffix":""},{"id":551234386,"identity":"31a6e765-0b9a-4d98-bbd1-7d876713a06c","order_by":3,"name":"Guangyan Wang","email":"","orcid":"","institution":"University of Texas Health Science Center","correspondingAuthor":false,"prefix":"","firstName":"Guangyan","middleName":"","lastName":"Wang","suffix":""},{"id":551234387,"identity":"a16d52e0-87e9-424e-882e-0ba1fe3236a6","order_by":4,"name":"Sumin Gu","email":"","orcid":"","institution":"UNIVERSITY OF TEXAS HEALTH SCIENCE CENTER AT SAN ANTONIO","correspondingAuthor":false,"prefix":"","firstName":"Sumin","middleName":"","lastName":"Gu","suffix":""}],"badges":[],"createdAt":"2025-08-31 15:50:12","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7501684/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7501684/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":98420991,"identity":"f5e87671-b83c-43e2-a1ac-dd1a4daee018","added_by":"auto","created_at":"2025-12-17 16:21:54","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2139385,"visible":true,"origin":"","legend":"Article File","description":"","filename":"msCDD.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7501684/v1_covered_25a0a3ff-e9b6-4367-9822-38c8b21bc783.pdf"},{"id":96877101,"identity":"4cb6a9f9-846b-46b4-9d04-802e0fa0fbdc","added_by":"auto","created_at":"2025-11-27 05:55:07","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":493151,"visible":true,"origin":"","legend":"Original Data Files","description":"","filename":"OriginalDatafiles.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7501684/v1/4275091983df2577f2ac4996.pdf"},{"id":96877100,"identity":"c9a8832b-83b7-42a4-bb70-2214f47619d0","added_by":"auto","created_at":"2025-11-27 05:55:07","extension":"pdf","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":2450130,"visible":true,"origin":"","legend":"Supplemental figures","description":"","filename":"Supplementaldata.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7501684/v1/2bd1ac283a3a67a8bcd83173.pdf"}],"financialInterests":"(Not answered)","formattedTitle":"Connexin Restrains EMT by Dual-Domain Mechanisms to Preserve Epithelial Identity","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 Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-7501684/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7501684/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"Epithelial–mesenchymal transition (EMT) contributes to fibrotic disease in multiple organs and underlies post-surgical complications such as posterior capsule opacification (PCO), the leading cause of vision loss following cataract surgery. The molecular mechanisms that preserve epithelial integrity and restrain EMT remain poorly defined. Connexins are conventionally regarded as gap junction proteins mediating intercellular communication, but here we identify connexin 50 (Cx50) as a previously unrecognized key regulator of EMT. Using in vitro, ex vivo, and in vivo chick and mouse models, we show that transforming growth factor-β2 (TGF-β2) downregulates Cx50 and induces EMT markers including α-smooth muscle actin and fibronectin, changes reversed by Cx50 overexpression. Mechanistically, the extracellular E2 domain of Cx50 mediates adhesive interactions that limit epithelial migration and EMT, while the C-terminal domain stabilizes β-catenin–E-cadherin complexes and prevents TGF-β2–driven E-cadherin loss and β-catenin nuclear translocation. Dominant-negative E2 mutants abolish protection, and Cx50 knockout mice exhibit accelerated EMT after extracapsular lens extraction. These findings redefine connexins beyond their classical role as gap junction proteins, establishing Cx50 as a dual-domain regulator of EMT and epithelial plasticity with implications for fibrotic responses in ocular and other epithelial tissues.","manuscriptTitle":"Connexin Restrains EMT by Dual-Domain Mechanisms to Preserve Epithelial Identity","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-11-27 05:55:02","doi":"10.21203/rs.3.rs-7501684/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"bde74287-ade7-42ee-b0d6-50e21168e6e1","owner":[],"postedDate":"November 27th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":58657229,"name":"Biological sciences/Cell biology/Mechanisms of disease"},{"id":58657230,"name":"Health sciences/Diseases/Trauma"}],"tags":[],"updatedAt":"2025-12-09T17:25:18+00:00","versionOfRecord":[],"versionCreatedAt":"2025-11-27 05:55:02","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7501684","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7501684","identity":"rs-7501684","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}