The autophagy genes ChATG4 and ChATG8 are required for reproductive development, virulence, and septin assembly in Cochliobolus heterostrophus

Autophagy is a highly conserved degrading process that is crucial for cell growth and development in eukaryotes, especially when they face starvation and stressful conditions. To evaluate the functions of Atg4 and Atg8 in mycelial growth, asexual and sexual development, and virulence in Cochliobolus heterostrophus, ΔChatg4 and ΔChatg8 mutants were generated by gene replacement. Strains deleted for ChATG4 and ChATG8 genes showed significant changes in vegetative growth and development of conidia and ascospores compared with the wild-type strain. The autophagy process was blocked and virulence was reduced dramatically in ΔChatg4 and ΔChatg8 mutants. In addition, deletion of ChATG4 and ChATG8 disordered Cdc10 subcellular localization and formation of septin rings. The direct physical interaction between ChAtg4 and ChAtg8 was detected by yeast two-hybrid assay, and ChAtg4-GFP was dispersed throughout the cytoplasm, although GFP-ChAtg8 appeared as punctate structures. All phenotypes were restored in complemented strains. Taken together, these findings indicate that ChATG4 and ChATG8 are crucial for autophagy to regulate fungal growth, development, virulence, and localization of septin in C. heterostrophus.