NSUN2 restrains renal fibrosis via m5C-stabilized GPX4 and ferroptosis inhibition.

BACKGROUND: Renal fibrosis (RFib) is a final common pathway in chronic kidney disease (CKD) progression. Although 5-methylcytosine (m5C) RNA modification can regulate mRNA fate, its role in RFib and redox-regulated cell death remains incompletely defined. METHODS: Public GEO cohorts were analyzed to identify dysregulated m5C regulators and enriched pathways in RFib. RFib was modeled in vivo using unilateral ureteral obstruction (UUO) and in vitro using TGF-β1-treated HK-2 cells. Gene set enrichment analysis (GSEA) and a ferroptosis-focused qPCR array were used to prioritize downstream programs. NSUN2 knockdown/overexpression experiments, dot blot assays, and m5C RNA immunoprecipitation followed by qPCR (MeRIP-qPCR) were performed to evaluate m5C-associated regulation of GPX4. Network pharmacology and in silico docking/dynamics were used to nominate candidate NSUN2-binding compounds. RESULTS: NSUN2 was consistently downregulated in fibrotic kidneys and TGF-β1-stimulated HK-2 cells, accompanied by ferroptosis activation. NSUN2 increased GPX4 protein abundance primarily by enhancing GPX4 mRNA stability, which correlated with increased m5C enrichment on GPX4 mRNA. Computational screening suggested that ginsenosides, particularly Rh2, may bind the NSUN2 catalytic pocket with favorable predicted affinity and complex stability. CONCLUSIONS: NSUN2 attenuates renal fibrogenesis by sustaining GPX4 expression through m5C-associated post-transcriptional regulation and thereby limiting ferroptosis. The identified ginsenosides provide testable leads for targeting the NSUN2-GPX4 axis, although further experimental validation is required.