{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T14:02:30Z","timestamp":1785420150081,"version":"3.56.0"},"posted":{"date-parts":[[2025,12,2]]},"group-title":"Genetics","reference-count":55,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,12,2]],"date-time":"2025-12-02T00:00:00Z","timestamp":1764633600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"award":["K08AR074529"],"award-info":[{"award-number":["K08AR074529"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["R01HD115266"],"award-info":[{"award-number":["R01HD115266"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["R01HD105033"],"award-info":[{"award-number":["R01HD105033"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["F32HD103376"],"award-info":[{"award-number":["F32HD103376"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["R01HL089221"],"award-info":[{"award-number":["R01HL089221"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["R01DK134408"],"award-info":[{"award-number":["R01DK134408"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["547486810"],"award-info":[{"award-number":["547486810"]}],"id":[{"id":"https:\/\/ror.org\/018mejw64","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,12,2]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Limb loss remains a significant clinical challenge, but regenerative medicine approaches such as gene therapy offer a promising strategy to trigger endogenous regeneration programs. Optimal vector configurations and molecular targets for appendicular skeletal repair are not well defined. Here, we leveraged insights from species with a high endogenous capacity for appendage regeneration to design an enhancer-directed gene delivery platform that functions during mouse digit regeneration, a well characterized model for partial limb regeneration in mammals. Single-cell RNA sequencing of zebrafish caudal fin regeneration, combined with expression data in regenerating salamander limbs and mouse digit tips, implicated the SP family of transcription factors as conserved, epidermally-expressed mediators of appendage regrowth. Null mutants of\n                  <jats:italic>Sp8<\/jats:italic>\n                  demonstrated impaired limb regeneration in salamanders, while conditional knockout of\n                  <jats:italic>Sp6<\/jats:italic>\n                  and\/or\n                  <jats:italic>Sp8<\/jats:italic>\n                  in the mouse basal epidermis resulted in defective bony digit tip regeneration, involving an IL-17 mediated osteoclastogenic program. Spatiotemporally focused expression of FGF8, a known target of SP factors, using a zebrafish-derived tissue regeneration enhancer element via adeno-associated viral vectors, could partially rescue digit tip regeneration in SP knockout mice and accelerate digit regeneration in wildtype mice. Our results demonstrate a contextual gene therapy approach to address limb loss based on genes like SP transcription factors conserved across multiple contexts of appendage regeneration.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Significance Statement<\/jats:title>\n                  <jats:p>\n                    Instructing regeneration of complex structures in mammals remains an unsolved problem. Gene therapy offers a compelling approach to foster endogenous regeneration by delivering therapeutic gene products to specific cells post injury. We identified a conserved regeneration-linked epidermal transcriptional program in mouse digit regeneration centered on the SP6 and SP8 transcription factors, involving inflammatory responses from osteoclasts. We engineered AAVs harboring a zebrafish tissue regeneration enhancer to direct FGF8 expression in the epidermis after amputation. This enhancer directed delivery partially rescued impaired digit regeneration in\n                    <jats:italic>Sp6<\/jats:italic>\n                    and\n                    <jats:italic>Sp8<\/jats:italic>\n                    conditional knockout mice and accelerated regrowth in wildtype digits. Our work links developmental signaling to adult regeneration and establishes a modular, injury site specific gene therapy framework that enables new interventions for limb healing.\n                  <\/jats:p>\n                <\/jats:sec>","DOI":"10.64898\/2025.12.01.691633","type":"posted-content","created":{"date-parts":[[2025,12,8]],"date-time":"2025-12-08T12:51:04Z","timestamp":1765198264000},"source":"Crossref","is-referenced-by-count":1,"title":["Enhancer-directed gene delivery for digit regeneration based on conserved epidermal factors"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0009-0005-9805-0632","authenticated-orcid":false,"given":"David A.","family":"Brown","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Katja 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