摘要Background Normal testicular development is essential for maintaining male fertility and reproductive performance in livestock.Leydig cells(LCs)play a central role in testicular physiology;however,the epigenetic mechanisms regulat-ing their development remain largely unclear.Methyltransferase-like 3(METTL3),a key m6A methylation enzyme,and microRNAs are increasingly recognised as critical regulators of this process.Results METTL3 expression in goat LCs markedly decreased during testicular development.This downregulation reduced m6A modification on pri-miR-145,impairing DiGeorge syndrome critical region 8-mediated processing and resulting in decreased levels of mature miR-145-3p.This reduction in miR-145-3p increased the expression of phosphoenolpyruvate carboxykinase 1(PCK1),which activated gluconeogenesis,increased intracellular glucose levels,and increased mitochondrial membrane potential.Consequently,this metabolic shift upregulated cell cycle-related genes(cyclin B1 and cyclin E2),promoting LC proliferation and testicular growth.Conclusions Our findings demonstrate that the METTL3/miR-145-3p/PCK1 axis is a key regulatory pathway linking epigenetic modification to the metabolic activity and proliferation of LCs.This mechanism provides novel insights into the molecular control of testicular development in male goats and may offer new targets for improving male reproductive capacity in livestock.