m ⁶ A demethylase CpALKBH regulates <i>CpZap1</i> mRNA stability to modulate the development and virulence of chestnut blight fungus

N⁶-methyladenosine (m⁶A) is the most abundant eukaryotic mRNA modification and is involved in various biological processes. Methyltransferases and demethylases regulate the m⁶A modification, but the regulatory role of m⁶A demethylases in fungi remains poorly understood. Here, we demonstrated that CpALKBH functions as a demethylase in <i>Cryphonectria parasitica</i>. The deletion of <i>CpALKBH</i> leads to a significant increase in m⁶A levels and a reduction in fungal growth, sporulation, and virulence. We identified <i>CpZap1</i> as a downstream target of CpALKBH, with CpALKBH regulating <i>CpZap1</i> mRNA stability in an m⁶A-dependent manner. Additionally, our findings indicate that methylation at position 1935A of <i>CpZap1</i> is regulated by both the CpALKBH demethylase and the CpMTA1 methyltransferase. Given its critical role in fungal development and virulence, overexpression of <i>CpZap1</i> can rescue abnormal phenotypes of ∆<i>CpALKBH</i> mutant. Overall, these findings contribute to improving our understanding of the role of m⁶A demethylase in fungi.