Abstract: Acid-producing fungal pathogens like Valsa mali enhance infectivity by secreting organic acids to acidify host environments, though the underlying cellular pH manipulation mechanisms remain unclear. Here, we identified VmAGP1 as a V. mali virulence factor whose knockout reduces virulence while heterologous expression in apples increases susceptibility. Using yeast two-hybrid (Y2H), bimolecular fluorescence complementation (BiFC), and co-immunoprecipitation (Co-IP) assays, we demonstrated that VmAGP1 interacts with apple receptor-like kinase MdLecRK2, which negatively regulates disease resistance. VmAGP1 promotes MdLecRK2 homo-dimerization, confirmed by luciferase complementation imaging (LCI) and Co-IP. Further studies reveal that MdLecRK2 interacts with and phosphorylates vacuolar H⁺-ATPase MdVHAc” 1, which also negatively regulates resistance. Flow cytometry shows that VmAGP1 expression lowers intracellular pH in apple protoplasts, further decreased by MdLecRK2/MdVHAc” 1 overexpression. We conclude that V. mali secretes VmAGP1 to induce MdLecRK2 homo-dimerization, triggering a phosphorylation cascade with MdVHAc” 1 that acidifies apple cells to facilitate infection. This study reveals a novel pH manipulation strategy in V. mali pathogenesis, identifying potential targets for controlling Apple Valsa canker.

Key words: acidi?cation, G‐type lectin receptor kinase, vacuolar H⁺‐ATPase, Vasla mali, VmAGP1