Functional Analysis Of Two Putative Rho GTPase Activating Proteins In Magnaporthe Grisea

Rho GTPases are implicated in the regulation of cellular processes as molecular switches that cycle between an active GTP-bound and an inactive GDP-bound state. Regulatory proteins termed Rho GAPs (Rho GTPase-activating proteins) stimulate the intrinsic GTPase activity and greatly enhance the GTP hydrolysis of the Rho GTP-binding proteins, thus causing their inactivation. RhoGAPs regulate the actin cytoskeleton organization and polarized cell growth, playing a critial role in morphogenesis.The rice blast fungus, Magnaporthe grisea, is taken as a model for the study of filamentous fungi. Its infection process involves a complex cell differentiation in response to host defense mechanism. We supposed that RhoGAPs participate in the regulation of this process. Based on the bioformation analysis, 8 putative RhoGAPs which contain the conserved RhoGAP domain were found in M. grisea. In this study, we have characterized two of the RhoGAPs (MgLrg1 and MgRga1) in M. grisea. We conducted the gene deletion of MgLRG1 and MgRGA1 through homologous recombination and analyzed the phenotype of Mglrg1 and Mgrga1 null mutants. The results showed that the growth rate of aerial hypha in Mglrg1 deletion mutant reduced dramatically and the mycelia branched more frequently at the ends. The conidiation ofâ–³Mglrg1 decreased remarkably and the normal conidia were defective in appressorium formation. Moreover, Mglrg1 deletion mutant lost pathogenicity completely. The deletion of Mgrga1 resulted in partially abnormal conidia and slight decrease of conidiation. The appressorium formation of Mgrga1 konckout mutant was delayed severely on hydrophobic surface, but it did not affect the development of mycelia and pathogenicity to plants. These data suggest that MgLrg1 is required for the regulation of the cell development, polarity growth and pathogenicity in M. grisea. Previous study has revealed that MgRga1 interacts directly with MgCdc42. MgRga1 may regulate the GTPase activity of MgCdc42 to control the cell polarity growth in M. grisea.