Trametinib ameliorated Adriamycin-induced podocyte injury by inhibiting METTL3 modified m6A RCAN1 RNA methylation

N6-methyladenosine (m⁶A) plays a crucial role in kidney diseases. Methyltransferase-like 3 (METTL3) as a key m⁶A writer can be regulated by trametinib. However, the epigenetic regulation of trametinib in focal segmental glomerulosclerosis (FSGS) remains unclear. We investigated whether trametinib protects podocytes by modulating METTL3-methylated target RNAs. Regulator of calcineurin 1 (RCAN1) was predicted as a target binding RNA of METTL3 by THEW database. Immunostaining of METTL3 and RCAN1 with podocyte marker Wilm's tumor-1 (WT-1) confirmed their localization within podocytes in renal biopsy from FSGS patients. Transfection METTL3 to human podocytes reduced WT-1, synaptopodin (SYNPO), and RCAN1 protein levels. Total m⁶A, m⁶A methylated RNA of RCAN1 increased and total RCAN1 mRNA decreased. Inhibition of METTL3 using siRNA or trametinib reversed these changes and attenuated the ADR-induced downregulation of WT-1 and SYNPO in vitro. In ADR-induced FSGS mice, trametinib ameliorated proteinuria, hypoalbuminemia, renal dysfunction, glomerulosclerosis and podocyte foot process effacement. Additionally, trametinib preserved podocyte function assessed by WT-1 and SYNPO as well as delayed renal fibrosis assessed by α-smooth muscle actin and fibronectin. Consistent with results in vitro, trametinib also decreased the ADR-induced upregulation of METTL3 and reversed the changed levels of total m⁶A, m⁶A methylated Rcan1 and total Rcan1 in FSGS mice. In conclusion, trametinib may serve as a renal protective agent for FSGS by regulating METTL3-dependent RCAN1 methylation levels.