Functional Analysis Of A Putative MgRhoGef1 Protein In Magnaporthe Oryzae

Rho1 GTPase is one of the most important Rho family members belonging to small GTP-binding proteins. It has two convertible forms: GDP-bound inactive and GTP-bound active forms. Guanine nucleotide releasing factor (also called GEF) regulated by an upstream signal interacts with GDP-bound form and catalyze the exchange of GDP for GTP to generate the active state of Rho GTPase. Based on bioinformatics analysis, the gene MGG₁2644 has a putative RhoGEF domain in its amino acid sequence, homologous to MgRhoGef1 in Saccharomyces cerevisiae. Phylogenetic analysis showed that the RhoGEF domain in MGG₁2644 are conserved but also showed divergent while the species evolved. Magnaporthe oryzae as an ideal model organism is used for our study. Knock-out mutants by homologous recombination grow slower than wild-type strain (WT), but the distance of two neighbor septa is nearly as normal, indicating that the MgRhoGef1 gene is related with cell division cycle. In addition, the mutants show lower ability to endure SDS than that of WT. The deletion mutants could not form conidia, but could develop appressoria at the hypha terminals, although there are fewer compared to the WT. These appressoria formed by deletion mutants lost the ability to infect the epidermal cell of onion as well as rice leaves, so we cannot see any lesions on the inoculated leaves. The complementary mutants recovered all these defects of deletion mutants indicating that the MgRhoGef1 is truly involved in these phenotypes. Based on the bioinformatics analysis, we infer that the MgRhoGef1 gene is the GEF of Rho1 protein. Yest two hybridization confirm that the interaction of MgRhoGef1 and its GEF domain with Rho1. Taking together, our study will be in favor of better understanding of Rho1 function, and further study would help to find an ideal target for controling the damage of Magnaporthe oryzae.