Palmitoylation induced activation of SMAD3 exacerbates colitis by promoting TRM cells differentiation

Palmitoylation induced activation of SMAD3 exacerbates colitis by promoting TRM cells differentiation | 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 Palmitoylation induced activation of SMAD3 exacerbates colitis by promoting TRM cells differentiation Yuejie Xu, Siqi Ji, Ping Jiang, Zhenguo Zhao, Qian Zhou, Dan Su, and 6 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7804610/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 04 Mar, 2026 Read the published version in Inflammation → Version 1 posted 11 You are reading this latest preprint version Abstract Tissue-resident memory T cells (TRM cells) have been shown to play an instrumental role in driving the onset and relapse of inflammatory bowel diseases (IBD). However, the underlying mechanism of TRM cells differentiation and its regulation in intestines remain to be unveiled. Mothers against decapentaplegic homolog 3 (SMAD3) is translocated from nucleus to membrane and activated in response to transforming growth factor beta (TGF-β), which is a key cytokine in the process of TRM cells polarization. Cysteine palmitoylation (S-palmitoylation) is a post-translational modification catalyzed by the DHHC family, regulating protein membrane associations. Genes associated with the classic SMAD3 signaling pathway, along with most genes in the DHHC family, were upregulated in TRM cells. Our study demonstrated that SMAD3 underwent reversible S-palmitoylation on Cys31 by DHHC6, leading to SMAD3 endomembrane recruitment and its subsequent colocalization with TGF-β receptor I (TGF-βRI) under TRM polarization conditions. The membrane recruitment of SMAD3 activated SMAD3 and subsequently upregulated the expression of its target genes, inducing the differentiation of TRM cells. In contrast, perturbation in DHHC6-induced palmitoylation with MYD-4 inhibited TRM cells differentiation and alleviated colitisin IBD model mice. Our work provides an example how the immune responses are regulated through the S-palmitoylation-dependent SMAD3 signaling in TRM cells differentiation and reveals protein S-palmitoylation as a potential target in IBD treatment, which could be of greater application considering the wide involvement of protein S-palmitoylation in the signal transduction in mammalian cells. Inflammatory bowel diseases S-palmitoylation TRM cells SMAD3 DHHC6 MYD-4 Full Text Additional Declarations No competing interests reported. Supplementary Files Supplementarymaterials.docx Cite Share Download PDF Status: Published Journal Publication published 04 Mar, 2026 Read the published version in Inflammation → Version 1 posted Editorial decision: Revision requested 20 Nov, 2025 Reviews received at journal 19 Nov, 2025 Reviews received at journal 04 Nov, 2025 Reviews received at journal 25 Oct, 2025 Reviewers agreed at journal 13 Oct, 2025 Reviewers agreed at journal 10 Oct, 2025 Reviewers agreed at journal 09 Oct, 2025 Reviewers invited by journal 09 Oct, 2025 Editor assigned by journal 08 Oct, 2025 Submission checks completed at journal 08 Oct, 2025 First submitted to journal 08 Oct, 2025 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-7804610","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":532018208,"identity":"56f7acbf-9b89-44af-baf5-934f8e0e7f12","order_by":0,"name":"Yuejie Xu","email":"","orcid":"","institution":"The Affiliated Wuxi People's Hospital of Nanjing Medical University, Wuxi People's Hospital","correspondingAuthor":false,"prefix":"","firstName":"Yuejie","middleName":"","lastName":"Xu","suffix":""},{"id":532018209,"identity":"e20f01e9-65b8-4fd6-94ff-31b12e0441f3","order_by":1,"name":"Siqi Ji","email":"","orcid":"","institution":"Shanghai Jiao 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differentiation","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"inflammation","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"ifla","sideBox":"Learn more about [Inflammation](https://www.springer.com/journal/10753)","snPcode":"10753","submissionUrl":"https://submission.nature.com/new-submission/10753/3","title":"Inflammation","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer 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However, the underlying mechanism of TRM cells differentiation and its regulation in intestines remain to be unveiled. Mothers against decapentaplegic homolog 3 (SMAD3) is translocated from nucleus to membrane and activated in response to transforming growth factor beta (TGF-β), which is a key cytokine in the process of TRM cells polarization. Cysteine palmitoylation (S-palmitoylation) is a post-translational modification catalyzed by the DHHC family, regulating protein membrane associations. Genes associated with the classic SMAD3 signaling pathway, along with most genes in the DHHC family, were upregulated in TRM cells. Our study demonstrated that SMAD3 underwent reversible S-palmitoylation on Cys31 by DHHC6, leading to SMAD3 endomembrane recruitment and its subsequent colocalization with TGF-β receptor I (TGF-βRI) under TRM polarization conditions. The membrane recruitment of SMAD3 activated SMAD3 and subsequently upregulated the expression of its target genes, inducing the differentiation of TRM cells. In contrast, perturbation in DHHC6-induced palmitoylation with MYD-4 inhibited TRM cells differentiation and alleviated colitisin IBD model mice. Our work provides an example how the immune responses are regulated through the S-palmitoylation-dependent SMAD3 signaling in TRM cells differentiation and reveals protein S-palmitoylation as a potential target in IBD treatment, which could be of greater application considering the wide involvement of protein S-palmitoylation in the signal transduction in mammalian cells.\u003c/p\u003e","manuscriptTitle":"Palmitoylation induced activation of SMAD3 exacerbates colitis by promoting TRM cells differentiation","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-10-23 14:11:40","doi":"10.21203/rs.3.rs-7804610/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-11-20T07:25:54+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-11-19T20:18:02+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-11-04T19:22:29+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-25T17:57:17+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"121522618139788310216664812444767704701","date":"2025-10-13T11:59:09+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"265443452248112056128693702944625787817","date":"2025-10-10T12:49:52+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"323814786628173970191934129104044791636","date":"2025-10-10T01:32:11+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-10-10T01:07:42+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-10-08T06:55:38+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-10-08T06:54:09+00:00","index":"","fulltext":""},{"type":"submitted","content":"Inflammation","date":"2025-10-08T06:16:45+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"inflammation","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"ifla","sideBox":"Learn more about [Inflammation](https://www.springer.com/journal/10753)","snPcode":"10753","submissionUrl":"https://submission.nature.com/new-submission/10753/3","title":"Inflammation","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"d93df89e-0a21-48ce-95c2-763c4b2cb03d","owner":[],"postedDate":"October 23rd, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-03-09T16:10:34+00:00","versionOfRecord":{"articleIdentity":"rs-7804610","link":"https://doi.org/10.1007/s10753-026-02466-x","journal":{"identity":"inflammation","isVorOnly":false,"title":"Inflammation"},"publishedOn":"2026-03-04 16:00:05","publishedOnDateReadable":"March 4th, 2026"},"versionCreatedAt":"2025-10-23 14:11:40","video":"","vorDoi":"10.1007/s10753-026-02466-x","vorDoiUrl":"https://doi.org/10.1007/s10753-026-02466-x","workflowStages":[]},"version":"v1","identity":"rs-7804610","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7804610","identity":"rs-7804610","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}