NAT10-dependent ac4C mRNA modification programs fibroblast pathogenicity in systemic sclerosis

Systemic sclerosis (SSc) is characterized by progressive fibrosis driven by persistent activation of dermal fibroblasts, yet the epigenetic and epitranscriptomic mechanisms sustaining fibroblast pathogenicity remain incompletely defined. Here, we investigated the role of N-acetyltransferase 10 (NAT10)-mediated N4-acetylcytidine (ac4C) RNA modification in fibroblast activation and fibrotic remodeling. Integrative analyses of single-cell transcriptomics, human SSc skin, in vitro assays, in vivo models, and fibroblast-specific knockout mice revealed that NAT10 is upregulated in pathogenic fibroblast populations and contributes to fibrotic progression. Genetic ablation or pharmacological inhibition of NAT10 attenuated fibroblast proliferation and extracellular matrix deposition across experimental systems. Multi-omics analyses identified COL11A1 and ZNF621 as downstream targets associated with NAT10, with site-specific ac4C modifications detected on both transcripts. Mechanistically, NAT10-dependent ac4C deposition was associated with increased mRNA stability and transcriptional output of these pro-fibrotic genes. Notably, suppression of ZNF621 partially reversed the pro-fibrotic phenotypes induced by NAT10 overexpression, supporting its functional relevance downstream. Together, these findings define an epitranscriptomic mechanism contributing to fibroblast activation in SSc and suggest that targeting the NAT10-ac4C axis may represent a potential therapeutic strategy for fibrotic disease. NAT10-dependent ac4C mRNA modification programs fibroblast pathogenicity in systemic sclerosis. N-acetyltransferase 10 (NAT10) regulates human dermal fibroblasts (HDF) proliferation and fibrosis via N4-acetylcytidine (ac4C) mRNA modification. During the fibrotic process in HDF, NAT10 expression is upregulated at both the transcriptional and translational levels. Upregulated NAT10 directly binds ZNF621 and COL11A1, increases the stability of their mRNAs in association with ac4C modification, and consequently promotes fibroblasts proliferation and fibrotic remodeling. Extracellular matrix (ECM); Remodelin, a small-molecule inhibitor commercially available for targeting NAT10. • NAT10 is aberrantly upregulated in systemic sclerosis fibroblasts and drives their pathological activation. • Genetic or pharmacological inhibition of NAT10 markedly suppresses fibroblast proliferation and tissue fibrosis in vivo. • NAT10 installs site-specific ac4C modifications on COL11A1 and ZNF621 mRNAs, enhancing their stability and pro-fibrotic output. • Disruption of the NAT10-ac4C-ZNF621 axis reverses fibrotic remodeling, identifying NAT10 as a druggable epitranscriptomic target in systemic sclerosis.