AaAO, AaPDE, and AaABC proteins promote Alternaria alternate f. sp. mali ALT7 invasion in apple leaves

Alternaria leaf spot, a fungal disease affecting apple production globally, incurs significant economic losses annually. The pathogenic fungus, A. alternata f. sp. mali (ALT), produces the host-specific AM-toxin and secretory proteins, which disrupt plant metabolism, leading to increased defoliation, fruit drop, compromised fruit quality, and reduced production. In this study, we isolated eight novel ALT strains from Fuji apple leaves exhibiting leaf spot. The most aggressive strain, ALT7, was identified. To determine ALT7 secretory proteins, LC-MS analysis was performed, which showed that three secreted proteins were detected: AaAO (alcohol oxidase), AaPDE (alkaline phosphatase), and AaABC (ATP-binding cassette transporter). Agrobacterium-mediated transgenesis confirmed the extracellular localization of GFP-fused AaAO, AaABC, and AaPDE. To investigate their function, we used fungal transgenesis. Overexpression of AaABC, AaAO, and AaPDE in ALT7 increased its pathogenicity. Conversely, knocking them down decreased ALT7 pathogenicity. Additionally, AaABC was found to facilitate the secretion of AaAO and AaPDE. And AaAO and AaPDE facilitate plant susceptibility by degrading plant cell walls, while AaABC plays a crucial role in their transport, thereby participating in the plant pathogenic process. In conclusion, our findings suggest that AaABC in A. alternata f. sp. mali mediates the secretion of toxic proteins AaAO and AaPDE.