METTL3-mediated m⁶A modification stabilizes COL1A1 mRNA to promote extracellular matrix accumulation in keloids

Keloids are pathological scars characterized by excessive collagen deposition and fibroblast hyperactivity, yet effective therapies remain limited due to incomplete understanding of their molecular drivers. Emerging evidence implicates N⁶-methyladenosine (m⁶A) RNA modification in fibrotic diseases, but its role in keloid pathogenesis is unclear. Here, we investigated the function and mechanism of the m⁶A methyltransferase METTL3 in keloid fibrosis. RNA sequencing of human keloid and normal skin tissues revealed significant upregulation of METTL3 and enrichment of extracellular matrix-related pathways. METTL3 was consistently overexpressed in keloid tissues and primary keloid fibroblasts (KF), which exhibited enhanced proliferation, migration, and resistance to apoptosis. Knockdown of METTL3 in KF suppressed these pro-fibrotic phenotypes and markedly reduced expression of α-SMA, TGF-β1, and collagen type I alpha 1 chain (COL1A1). Mechanistically, METTL3 deposited m⁶A modifications on the coding sequence of COL1A1 mRNA, enhancing its stability. YTHDF1 recognized these m⁶A marks and promoted COL1A1 mRNA translation. RNA pull-down and gene-specific m⁶A-PCR confirmed direct binding of YTHDF1 to m⁶A-modified COL1A1; loss of YTHDF1 reduced COL1A1 protein levels without affecting its mRNA abundance. Rescue experiments in which COL1A1 overexpression restored the fibrotic phenotypes suppressed by METTL3 knockdown established COL1A1 as a critical downstream effector of METTL3. Importantly, intradermal delivery of METTL3 shRNA in a rat tension-driven excisional scar model significantly attenuated scar formation, collagen accumulation, and fibrotic marker expression. Our findings establish METTL3 as a central epitranscriptomic regulator of keloid fibrosis by stabilizing COL1A1 mRNA and enabling YTHDF1-mediated translational enhancement, forming an m⁶A-YTHDF1-COL1A1 axis, highlighting its potential as a novel therapeutic target for pathological scarring.