METTL16-m6A-PGC-1α axis contributes to exercise-induced mitochondrial adaptations in skeletal muscle of high-fat diet-fed insulin-resistant mice

The dynamic change of N6-methyladenosine (m 6 A) modification on substrate RNA molecules plays a critical role in different biological processes and disease pathogenesis. Although the beneficial effects of exercise training (ET) on skeletal muscle insulin resistance (IR) are well-established, the contribution of RNA m 6 A modification in ET-related adaptations in high-fat diet (HFD)-induced IR remains unclear. In this study, we show that exercise stimulation triggers a dynamic shift in skeletal muscle m 6 A modification levels during HFD consumption. As a key m 6 A methyltransferase, METTL16 was downregulated in HFD-fed mice and upregulated by ET at both the mRNA and protein levels. In vitro, METTL16 knockdown disrupted mitochondrial ultrastructure, reduced electron transport chain complex activities, and decreased the NAD + /NADH ratio, ATP content, and mitochondrial membrane potential, indicating impaired mitochondrial function. Concomitantly, METTL16 loss lowered m 6 A on PGC-1α mRNA, reducing its stability and protein abundance and blunting insulin signalling, whereas PGC-1α overexpression partially reversed these defects. In conclusion, METTL16 functions as an exercise-responsive m 6 A methyltransferase that may modulate PGC-1α, mitochondrial function, and insulin-related signalling in HFD skeletal muscle, implicating the METTL16-m 6 A-PGC-1α axis in exercise-induced metabolic adaptations.