Wheat (Triticum aestivum L.) is a staple food of more than 50% of global population. Rhizoctonia cerealis is the causal agent of sharp eyespot, a devastating disease of cereal crops including wheat. Cutinases produced by fungal pathogens play important roles in host-pathogen compatible interactions, but little is known about cutinases in R. cerealis. In this study, we identified a total of six
cutinase encoding genes from R. cerealis genome, designated as RcCUT1-RcCUT6, analyzed their expression patterns during the
infection, and determined virulence role for RcCUT1. All the
proteins, RcCUT1-RcCUT6, contain a highly conserved GYSKG motif and another conserved C-x(3)-D-x(2)-C-x(2)-[GS]-[
GSD]-x(4)-[AP]-H motif in the
carbohydrate esterase 5 domain. The RcCUT1, RcCUT2, RcCUT4, and RcCUT5 are predicted to be secreted
proteins containing four
cysteine residues. These six
cutinase genes had different expression patterns during the
fungal infection process to wheat, among which RcCUT1 was highly expressed across all the
infection time points but RcCUT6 was not expressed at all and the others were expressed only at certain time points. Further, RcCUT1 was heterologously expressed in Escherichia coli to obtain a purified
protein. The purified RcCUT1 was shown to possess the
cutinase activity and be able to induce
necrosis, H2O2 accumulation, and expression of defense-related genes when infiltrated into wheat and Nicotiana benthamiana leaves. In contrast, RcCUT1
protein with
serine mutation at the first motif had no
cutinase activity, consequently lost the ability to induce
necrosis. Noticeably, application of the purified RcCUT1 with R. cerealis led to significantly higher levels of the disease in wheat leaves than application of the fungus alone. These results strongly suggest that RcCUT1 serves as a
virulence factor for the fungus. This is the first investigation of the
cutinase genes in R. cerealis and the findings provide an important insight into pathogenesis mechanisms of R. cerealis on wheat.