Functional Analysis Of Three Putative ARF Proteins In Magnaporthe Grisea

ADP Ribosylation Factors （ARFs） are members of the ARF family of20kDa GTP-binding proteins of the Ras superfamily. They were originally identified as the protein cofactors required for efficient in vitro ADP ribosylation of the stimulatory regulatory component of adenylate cyclase, Gas, by cholera toxin. Recently it has been shown that ARF plays a critical role in vesicular traffic and actin remodeling.In this study, we want to know the function of ARF in Magnaporthe grisea, so we used Magnaporthe grisea database and bioinformatic softwares to compare the homology of deduced ARF in Magnaporthe grisea. Then we constructed three knockout vectors of MGG01574、MGG04976and MGG08859, two over-expression vectors of MGG₀1574and MGG₀4976. Through protoplast transformation, we obtained the correspondent arf knockout mutants and ARF overexpression strains. Phenotype analysis showed that the knockout mutants of MGG₀1574and MGG₀4976have no phenotype changes in comparison with the wild type strain, but mycelia growth of the knockout mutants of MGG₀8859reduced dramatically. And overexpression of MGG₀1574caused the mycelia growth reduced slightly, the sporualtion of overexpression of MGG₀1574and MGG₀4976decreased in comparison with GUY11. Conidium germination, appressoria formation and pathogenicity were not affected in all mutants tested. Taking together, it’s indicated that these putative ARFs participate in mycelia growth and conidiation, however, they are not essential, the dynamics of ARFs may be important, therefore overexpression of ARFs inhibits the mycelial growth on the contrary.