Structural basis of Fusarium myosin I inhibition by phenamacril.

Fusarium is a genus of filamentous fungi that includes species that cause devastating diseases in major staple crops, such as wheat, maize, rice, and barley, resulting in severe yield losses and mycotoxin contamination of infected grains. Phenamacril is a novel fungicide that is considered environmentally benign due to its exceptional specificity; it inhibits the ATPase activity of the sole class I myosin of only a subset of Fusarium species including the major plant pathogens F. graminearum, F. asiaticum and F. fujikuroi. To understand the underlying mechanisms of inhibition, species specificity, and resistance mutations, we have determined the crystal structure of phenamacril-bound F. graminearum myosin I. Phenamacril binds in the actin-binding cleft in a new allosteric pocket that contains the central residue of the regulatory Switch 2 loop and that is collapsed in the structure of a myosin with closed actin-binding cleft, suggesting that pocket occupancy blocks cleft closure. We have further identified a single, transferable phenamacril-binding residue found exclusively in phenamacril-sensitive myosins to confer phenamacril selectivity.

Regulatory roles of introns in fungicide sensitivity of Fusarium graminearum.

Although the roles of introns have been much debated in eukaryotic organisms, none of them have been functionally characterized in Fusarium graminearum. In this study, we characterized the roles of introns in regulation of fungicide-sensitivity of F. graminearum. ß2 tub, cyp51A and myosin-5 are important target genes of benzimidazoles, triazoles and cyanoacrylates respectively. To explore the sensitivity regulation functions of introns in target genes, several detailed deletion studies were completed on the intronic regions of ß2 tub, cyp51A and myosin-5. Phenotypic analyses showed that deletion of the fourth intron from ß2 tub gene (designated ß2 Δi4), the sole intron from cyp51A gene (cyp51A-Δi) and the second intron from myosin-5 gene (myo5-Δi2) exhibited an increased sensitivity to corresponding fungicides. In contrast, deletion of the first or second intron from ß2 tub gene exhibited a decreased sensitivity to carbendazim. qRT-PCR showed that the mRNA transcript levels of target genes were significantly downregulated in ß2 Δi4, cyp51A-Δi and myo5-Δi2 respectively. Meanwhile, Western blot assays revealed that the protein expression levels of ß2 tub was also dramatically reduced in ß2 Δi4, but accumulated in ß2 Δi1 and ß2 Δi2. Overall, our results indicate that introns in target genes significantly regulate the fungicide-sensitivity by influencing expression of the corresponding resident genes in F. graminearum.