METTL3/YTHDF1-mediated m6A modification stabilizes USP12 to deubiquitinate FOXO3 and promote apoptosis in sepsis-induced myocardial dysfunction

Sepsis-induced myocardial dysfunction (SIMD) is a life-threatening complication primarily driven by inflammation, yet its molecular mechanisms remain unclear. In this study, we identified significant upregulation of the m⁶A methyltransferase METTL3 (methyltransferase-like 3), the m⁶A reader protein YTHDF1 (YTH N6-methyladenosine RNA binding protein 1), as well as increased expression levels of USP12 (ubiquitin-specific peptidase 12), FOXO3 (forkhead box O3), and key molecules in the intrinsic apoptotic pathway, PUMA (p53 upregulated modulator of apoptosis) and BAX (Bcl-2-associated X), through proteomic profiling in an LPS (Lipopolysaccharide)-induced SIMD mouse model. In vitro and in vivo experiments demonstrated that METTL3 and YTHDF1 regulated USP12 mRNA expression and stability through m⁶A modification. Elevated USP12 interacted with FOXO3, preventing its ubiquitin-mediated degradation, which enhanced FOXO3 binding to the PUMA promoter, leading to upregulation of PUMA. PUMA upregulation initiated the intrinsic apoptotic pathway, activating downstream BAX, Apaf1 (apoptotic protease-activating factor 1), and Caspases, ultimately driving SIMD. Inhibition of METTL3 (with STM2457), YTHDF1 (with Ebselen), or PUMA (with CLZ-8) significantly suppressed intrinsic apoptosis and alleviated SIMD symptoms. These findings underscore the critical role of METTL3/YTHDF1-dependent m⁶A modification in modulating the USP12-FOXO3-PUMA-BAX-Apaf1-Caspases signaling axis in SIMD, and suggest that targeting this pathway may offer a potential therapeutic strategy for SIMD.