Multi-omics analysis reveals NSUN2-mediated m5C methylation of DOCK9 regulating vascular stability in brain arteriovenous malformations

RNA 5-methylcytosine (m5C) methylation is a key post-transcriptional modification, yet its role in vascular stability remains unclear. Through integrative m5C-RNA immunoprecipitation and transcriptomic, proteomic, and single-cell analyses of ruptured and unruptured human brain arteriovenous malformations (bAVMs), we find DOCK9 to be an endothelial-specific, m5C-regulated gene downregulated in the ruptured group. Functional assays in endothelial cells and CRISPR-Cas9 zebrafish models show that DOCK9 controls endothelial proliferation, migration, and vascular integrity. Endothelial-specific overexpression of dock9 rescues vascular defects in dock9-deficient zebrafish. Mechanistically, the RNA methyltransferase NSUN2 directly binds and methylates DOCK9 mRNA to stabilize its expression. NSUN2 knockdown mimics dock9 deficiency in zebrafish, supporting a functional NSUN2-DOCK9 axis. These findings show a molecular pathway linking m5C dysregulation to cerebrovascular instability and bAVM rupture. Targeting this axis may offer a strategy for stabilizing fragile cerebrovascular lesions.