Epigenetic signature of human vitamin D3 and IL-10-conditioned regulatory DCs 

Epigenetic signature of human vitamin D3 and IL-10-conditioned regulatory DCs | 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 Epigenetic signature of human vitamin D3 and IL-10-conditioned regulatory DCs Silvia Liu, Alan F. Zahorchak, Steven F. Dobrowolski, Diana M. Metes, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4385501/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 20 Nov, 2024 Read the published version in Scientific Reports → Version 1 posted 12 You are reading this latest preprint version Abstract During differentiation of precursor cells into their destination cell type, cell-fate decisions are enforced by a broad array of epigenetic modifications, including DNA methylation, which is reflected by the transcriptome. Thus, regulatory dendritic cells (DCregs) acquire specific epigenetic programs and immunomodulatory functions during their differentiation from monocytes. To define the epigenetic signature of human DCregs generated in vitamin D3 (vitD3) and IL-10 compared to immune stimulatory DCs (sDCs), we measured levels of DNA methylation by whole genome bisulfite sequencing (WGBS). Distinct DNA methylation patterns were acquired by DCregs compared to sDCs. These patterns were located mainly in transcriptional regulatory regions. Associated genes were enriched in STAT3-signaling and valine catabolism in DCregs; conversely, pro-inflammatory pathways, e.g. pattern recognition receptor signaling, were enriched in sDCs. Further, DCreg differentially- methylated regions (DMRs) were enriched in binding motifs specific to the immunomodulatory transcription factor Krueppel-like factor 11 (KLF11), while activator protein-1 (AP-1) (Fos:Jun) transcription factor-binding motifs were enriched in sDC DMRs. Using publicly-available data-sets, we defined a common epigenetic signature shared between DCregs generated in vitD3 and IL-10, or dexamethasone or vitD3 alone. These insights may help pave the way for design of epigenetic-based approaches to enhance the production of DCregs as effective therapeutic agents. Figures Figure 1 Figure 2 Figure 3 Full Text Additional Declarations No competing interests reported. Supplementary Files Liuetal.SciReportsSuppl.Figure052024.pdf Liuetal.SciReportsSuppl.text052024copy.pdf Suppl.Table1.xlsx Suppl.Table2.xlsx Cite Share Download PDF Status: Published Journal Publication published 20 Nov, 2024 Read the published version in Scientific Reports → Version 1 posted Editorial decision: Revision requested 27 Sep, 2024 Reviews received at journal 18 Sep, 2024 Reviewers agreed at journal 01 Sep, 2024 Reviewers agreed at journal 24 Jul, 2024 Reviews received at journal 17 Jun, 2024 Reviewers agreed at journal 10 Jun, 2024 Reviewers agreed at journal 07 Jun, 2024 Reviewers invited by journal 24 May, 2024 Editor assigned by journal 24 May, 2024 Editor invited by journal 15 May, 2024 Submission checks completed at journal 13 May, 2024 First submitted to journal 07 May, 2024 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. 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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-4385501","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":305141503,"identity":"b908403e-22c7-422c-b032-acf196eaaac0","order_by":0,"name":"Silvia Liu","email":"","orcid":"","institution":"University of Pittsburgh","correspondingAuthor":false,"prefix":"","firstName":"Silvia","middleName":"","lastName":"Liu","suffix":""},{"id":305141504,"identity":"a41fe3b5-c883-4fa8-adc0-a9425c7b9373","order_by":1,"name":"Alan F. Zahorchak","email":"","orcid":"","institution":"University of Pittsburgh","correspondingAuthor":false,"prefix":"","firstName":"Alan","middleName":"F.","lastName":"Zahorchak","suffix":""},{"id":305141505,"identity":"bc21d065-6652-4f70-9ab8-2e3d1c5b22b0","order_by":2,"name":"Steven F. Dobrowolski","email":"","orcid":"","institution":"University of Pittsburgh","correspondingAuthor":false,"prefix":"","firstName":"Steven","middleName":"F.","lastName":"Dobrowolski","suffix":""},{"id":305141506,"identity":"c89af1a7-83b4-47e5-9211-4f71ed264e85","order_by":3,"name":"Diana M. Metes","email":"","orcid":"","institution":"University of Pittsburgh","correspondingAuthor":false,"prefix":"","firstName":"Diana","middleName":"M.","lastName":"Metes","suffix":""},{"id":305141507,"identity":"4197a214-c216-4885-a48e-8f0a72164760","order_by":4,"name":"Hossam A. Abdelsamed","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABAElEQVRIiWNgGAWjYBACAwYeBgmGAjDb8AFCnI2QFgMw29gAoZpILWYSRGkxZz978MYHgzp53fbmbRU/22oT589vfsDwoewwTi2WPXnJljMMDhtuO3Os7GZv2/HEDcfYDBhnnMOtxeBAjpk0j8EBxm03csxu8LYdS9zAxmDAzNuGR8v5N2bSfwzq7Lfdf2NW+BeoZX4b+wfmv/i0AA2XBhqbuO0GjxnQ8JrEhmM8BsyMeLW8S7bsMTicvO1MWrG0zLkDxhuO5RQc7DmXjsdhuQdv/Kios912/PDGj2/K6mTnNx/f+OBHmTVOLegA4p4DRKsHgjpSFI+CUTAKRsEIAQCR5l8NMiQklQAAAABJRU5ErkJggg==","orcid":"","institution":"Augusta University","correspondingAuthor":true,"prefix":"","firstName":"Hossam","middleName":"A.","lastName":"Abdelsamed","suffix":""},{"id":305141508,"identity":"496be9c3-0381-4dba-afec-98948388fdd0","order_by":5,"name":"Angus Thomson","email":"","orcid":"","institution":"University of Pittsburgh","correspondingAuthor":false,"prefix":"","firstName":"Angus","middleName":"","lastName":"Thomson","suffix":""}],"badges":[],"createdAt":"2024-05-07 22:56:09","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4385501/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4385501/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1038/s41598-024-79299-x","type":"published","date":"2024-11-20T15:57:32+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":56964916,"identity":"8818c45e-6ba6-48f7-ae13-006950ef9521","added_by":"auto","created_at":"2024-05-22 19:32:33","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":161444,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003e\u003cstrong\u003eGenome-wide changes in DNA methylation of human DCregs compared with sDCs;\u003c/strong\u003e\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(A) \u003c/strong\u003eIn vitro culture schema showing the generation of DCregs and sDCs from human peripheral blood monocytes; \u003cstrong\u003e(B) \u003c/strong\u003eSchematic overview showing DNA methylation computational analyses pipeline for DCregs vs sDCs; \u003cstrong\u003e(C) \u003c/strong\u003ePrincipal component analyses (PCA) of differentially-methylated regions (DMRs) in DCregs and sDCs; \u003cstrong\u003e(D) \u003c/strong\u003eDNA methylation heat map showing two clusters (A and B) representing levels of methylation in DMRs (DCregs vs sDCs) represented as normalized methylation levels (Z-score -2 to 2); \u003cstrong\u003e(E) \u003c/strong\u003eVenn diagram showing the common and distinct pathways in DCregs and sDCs using the genes annotated to DMRs within introns, exons, 5’ distal regions, intergenic, promoter, and other regions (other = 3UTR, 5UTR, 3’ distal, downstream) as an input in ingenuity pathway analyses (IPA); \u003cstrong\u003e(F) \u003c/strong\u003ethe nature of the pathways enriched in DCregs and sDCs using IPA.\u003c/p\u003e","description":"","filename":"Liuetal.SciReportsMainFigures0520241.png","url":"https://assets-eu.researchsquare.com/files/rs-4385501/v1/5ac5c8b2c46dd9f2b6004a99.png"},{"id":56964919,"identity":"7bd51bbf-1633-4a0e-99ae-8782e7f7b8ce","added_by":"auto","created_at":"2024-05-22 19:32:34","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":314764,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003e\u003cstrong\u003eNature, number and distribution of DMRs comparing DCregs and sDCs \u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e(A) \u003c/strong\u003eNumber and distribution of the DMRs in DCregs and sDCs (other = 3UTR, 5UTR, 3’ distal, downstream); \u003cstrong\u003e(B) \u003c/strong\u003eVenn diagram showing the common and distinct genes annotated to regulatory gene transcription regions including 5’ distal regions and promoters in DCregs and sDCs; \u003cstrong\u003e(C) \u003c/strong\u003ethe nature of the pathways enriched in DCregs and sDCs using genes annotated in B; \u003cstrong\u003e(D) \u003c/strong\u003eUpstream transcription factors (TF) and regulators associated with genes annotated to regulatory transcription regions analyzed in B; \u003cstrong\u003e(E) \u003c/strong\u003eVenn diagram showing common and distinct discovered TF motifs within DMRs in DCregs vs sDCs \u003cstrong\u003e(F) \u003c/strong\u003eMotif discovery analyses showing enrichment of Krueppel-like factor 11 (KLF11) motif in unmethylated DMR of DCregs (upper panel) as well as activator protein-1 (AP1) TF motif enriched in unmethylated DMRs of sDCs (lower panel).\u003c/p\u003e","description":"","filename":"Liuetal.SciReportsMainFigures0520242.png","url":"https://assets-eu.researchsquare.com/files/rs-4385501/v1/2eb2e91f551958530ed69575.png"},{"id":56964921,"identity":"6cbc1332-5837-4d5d-8c9b-babf2620d15c","added_by":"auto","created_at":"2024-05-22 19:32:34","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":443596,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003e\u003cstrong\u003eDCs generated in vitD3 and IL-10 exhibit a tolerogenic transcriptional and epigenetic signature \u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e(A) \u003c/strong\u003eVenn diagram showing distinct and overlapping unmethylated DMRs in DCregs generated using three different culture conditions (VitD3 alone, VitD3/IL-10, and Dexamethasone alone); \u003cstrong\u003e(B) \u003c/strong\u003eSummary table showing common core epigenetic signature list between the three DCreg culture conditions \u003cstrong\u003e(C) \u003c/strong\u003eHistograms and bar graphs showing expression of CXCR4 (incidence of positive cells and mean fluorescence intensity (MFI)) among CD11c+ HLA-DR+ DCs (DCregs vs sDCs); *p\u0026lt;0.05 unpaired parametric t-test with Welch’s correction; \u003cstrong\u003e(D) \u003c/strong\u003escatter plots showing the nature and correlation of the genes between methylation differences (DCregs-sDCs) and gene expression changes (Log2 DCregs-sDCs). Pearson’s correlation (r2 =-0.83) was used by comparing highly methylated genes with low gene expression in the same set of genes; \u003cstrong\u003e(E) \u003c/strong\u003eBar graph showing levels of methylation across CD163 promoter region (Chr12: 7665530-766596) in DCregs compared sDCs in six different healthy adult donors (top panel), Unpaired non-parametric t-test (Mann-Whitney test) *p\u0026lt;0.05 and paired analyses showing mean fluorescence intensity (MFI)) on CD11c+ HLA-DR+ DCs (DCregs vs sDCs) [bottom panel]. Non-parametric paired t-test (Wilcoxon matched-pairs signed rank test *p\u0026lt;0.05).\u003c/p\u003e","description":"","filename":"Liuetal.SciReportsMainFigures0520243.png","url":"https://assets-eu.researchsquare.com/files/rs-4385501/v1/a7fbbac03063ff9a77103238.png"},{"id":69834892,"identity":"348207ab-4b6d-41d1-8282-585f4fc9d652","added_by":"auto","created_at":"2024-11-25 16:10:05","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1139177,"visible":true,"origin":"","legend":"","description":"","filename":"Liuetal.SciReportsMaintext052024.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4385501/v1_covered_95774628-0f2b-4b49-a72b-6f85daee8540.pdf"},{"id":56964920,"identity":"45185ff0-4beb-48d3-9c5a-b5a2f7a74805","added_by":"auto","created_at":"2024-05-22 19:32:34","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":591607,"visible":true,"origin":"","legend":"","description":"","filename":"Liuetal.SciReportsSuppl.Figure052024.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4385501/v1/6cd32f4811a7634c6512a43d.pdf"},{"id":56964918,"identity":"33cc0de1-de55-4568-93f8-9b77a34f4a29","added_by":"auto","created_at":"2024-05-22 19:32:34","extension":"pdf","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":48215,"visible":true,"origin":"","legend":"","description":"","filename":"Liuetal.SciReportsSuppl.text052024copy.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4385501/v1/9ba9efc666de9842a87c2a75.pdf"},{"id":56964922,"identity":"8a36b89b-6596-4c79-8dd1-a3c213f1f4f4","added_by":"auto","created_at":"2024-05-22 19:32:34","extension":"xlsx","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":79614,"visible":true,"origin":"","legend":"","description":"","filename":"Suppl.Table1.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-4385501/v1/67d7b66434e809748454bcbb.xlsx"},{"id":56964923,"identity":"1fbaeb4e-1408-4b90-bdc2-b1be93e31fc8","added_by":"auto","created_at":"2024-05-22 19:32:35","extension":"xlsx","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":1237894,"visible":true,"origin":"","legend":"","description":"","filename":"Suppl.Table2.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-4385501/v1/1d675548d99e3efe545a1c85.xlsx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Epigenetic signature of human vitamin D3 and IL-10-conditioned regulatory DCs ","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":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-4385501/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4385501/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"During differentiation of precursor cells into their destination cell type, cell-fate\ndecisions are enforced by a broad array of epigenetic modifications, including DNA methylation, which is reflected by the transcriptome. Thus, regulatory dendritic cells (DCregs) acquire specific epigenetic\nprograms and immunomodulatory functions during their differentiation from monocytes. To define the\nepigenetic signature of human DCregs generated in vitamin D3 (vitD3) and IL-10 compared to immune\n stimulatory DCs (sDCs), we measured levels of DNA methylation by whole genome bisulfite\nsequencing (WGBS). Distinct DNA methylation patterns were acquired by DCregs compared to sDCs.\nThese patterns were located mainly in transcriptional regulatory regions. Associated genes were\n enriched in STAT3-signaling and valine catabolism in DCregs; conversely, pro-inflammatory pathways,\n e.g. pattern recognition receptor signaling, were enriched in sDCs. Further, DCreg differentially-\n methylated regions (DMRs) were enriched in binding motifs specific to the immunomodulatory\n transcription factor Krueppel-like factor 11 (KLF11), while activator protein-1 (AP-1) (Fos:Jun)\n transcription factor-binding motifs were enriched in sDC DMRs. Using publicly-available data-sets, we\n defined a common epigenetic signature shared between DCregs generated in vitD3 and IL-10, or\n dexamethasone or vitD3 alone. These insights may help pave the way for design of epigenetic-based\n approaches to enhance the production of DCregs as effective therapeutic agents.","manuscriptTitle":"Epigenetic signature of human vitamin D3 and IL-10-conditioned regulatory DCs ","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-05-22 19:32:29","doi":"10.21203/rs.3.rs-4385501/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-09-27T04:50:59+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-09-18T18:04:54+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"58396596242265105981445529474589481540","date":"2024-09-01T23:43:21+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"184435695860346910602467749162940219126","date":"2024-07-24T06:12:18+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-06-17T09:04:20+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"117180360429239881346139824839119686714","date":"2024-06-10T13:20:35+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"299305856196950295661941723576855149889","date":"2024-06-07T10:09:58+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-05-24T06:35:56+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-05-24T06:10:39+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2024-05-15T16:08:01+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-05-14T03:54:00+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2024-05-07T22:54:42+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"d888b8c5-94aa-4344-819c-10a6ad4ff603","owner":[],"postedDate":"May 22nd, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2024-11-25T16:02:07+00:00","versionOfRecord":{"articleIdentity":"rs-4385501","link":"https://doi.org/10.1038/s41598-024-79299-x","journal":{"identity":"scientific-reports","isVorOnly":false,"title":"Scientific Reports"},"publishedOn":"2024-11-20 15:57:32","publishedOnDateReadable":"November 20th, 2024"},"versionCreatedAt":"2024-05-22 19:32:29","video":"","vorDoi":"10.1038/s41598-024-79299-x","vorDoiUrl":"https://doi.org/10.1038/s41598-024-79299-x","workflowStages":[]},"version":"v1","identity":"rs-4385501","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4385501","identity":"rs-4385501","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}