The Phosphorylation in TGY Motif of the Mitogen-Activated Protein Kinase Hog1 Governs Fludioxonil Resistance and Biological Characteristics via Its Subcellular Localization in Filamentous Fungi.

Fludioxonil is a widely used phenylpyrrole fungicide. Previously, fludioxonil resistance was primarily associated with mutations in the histidine kinase Os1. However, highly resistant isolates without such mutations suggest the existence of additional mechanisms. This study identified a novel resistance pathway centered on the TGY motif of the terminal MAPK FgHog1 in Fusarium graminearum. Site-directed substitutions of phosphorylatable threonine and tyrosine residues in the TGY motif generated mutants FgHog1-T171G, FgHog1-Y173A, and double mutant FgHog1-T171G&Y173A. All these mutations abolished detectable FgHog1 phosphorylation and conferred extremely high fludioxonil resistance. Loss of FgHog1 phosphorylation also blocked fludioxonil-induced nuclear translocation of FgHog1. However, nonphosphorylatable mutants exhibited severe defects in vegetative growth, sporulation, virulence, glycerol accumulation, acetic acid utilization, and sexual development, demonstrating strong pleiotropy. These results indicate that FgHog1 phosphorylation governs fludioxonil sensitivity and multiple biological functions, while the associated fitness costs may limit resistance development in field populations.