Abstract / Summary
Abstract Tissue repair requires rapid changes in cell state and gene function, yet most studies have focused on overall gene expression and have not resolved how alternative transcript isoforms contribute to repair. Here, we generated a cell-type-specific, isoform-resolved atlas of human skin wound healing by integrating PacBio Iso-Seq, deep short-read RNA sequencing, and single-cell transcriptomics in a longitudinal human in vivo wound model with donor-matched intact skin and wound tissues. PacBio Iso-Seq established a high-confidence full-length transcriptome reference for fibroblasts and keratinocytes, revealing marked cell-type specificity among previously unannotated transcripts. Quantitative profiling of day-7 wound cells uncovered extensive differential isoform expression and isoform switching beyond changes in overall gene expression. Alternative splicing was dominated by skipped-exon events, with FN1 emerging as a major wound-regulated target in fibroblasts. Functional interrogation of wound-regulated LDLR and TNFRSF21 isoforms revealed distinct effects on lipoprotein uptake, inflammatory responses, proliferation, and migration. Single-cell exon-skipping analysis further resolved wound-associated, cell-state-specific splicing programs. Wounding also induced widespread changes in alternative promoter usage. Single-cell alternative polyadenylation analysis revealed dynamic wound-associated 3′ end regulation, including pronounced 3′ untranslated region shortening in proliferative fibroblasts. Together with an interactive public browser, our study establishes transcript isoform remodeling as a key regulatory dimension of human skin wound healing and provides a resource for isoform-aware studies of tissue repair.