A Novel Zn-finger Protein As A Virulence Factor In Magnaporthe Grisea

Rice blast disease caused by Magnaporthe grisea is one of the most important diseases of rice. Rice blast fungus has become a model organism to investigate the interactions between plants and plant-pathogens fungi. The study on the pathogenicity at molecular level is on the behalf of understanding the mechanism of pathogenicity of M. grisea, the interaction between rice blast fungus and rice, providing more potential targets for the design of novel, environmental safe compounds. The thesis is about the isolation of targeted gene on pathogenicity and the gene function.A mutant H680 with dramatically reduced pathogenicity against the susceptible rice cultivar Tsuyuake was obtained through screening REMI transformants library. The number of lesions is 5% of that of wild type P131, and the growth, percentage of germination, appressorium and penetration peg formation on onion epidermis are reduced to some extents. After 48h of inoculation, percentage of appressorim formation is not increased. 60% of germ tube cannot develop into appressorium, which are going on vegetative growth The result of sexual hybridization and Southern blot analysis show that the phenotype of the mutant cosegragated with insertion marker. There is just one integration site and single copy of insertion plasmid in the mutant H680 genome.The target gene was cloned through plasmid rescue. The flanking sequence showed the linearized pUCATPH insertion occurred in the site of 19086bp on the contg2.1287 by the search of rice blast fungus genomic database The insertion was predicted to lie between two genes by GENSCAN analysis, 171bp and 233bp away from the translation initial site ATG respectively. One gene has 987bp with no intron encodes a protein of 328aa; the other encodes a protein of 1115aa with two extrons.It was proved that the smaller one was tagged and is responsible for the appressorium formation and the pathogenicity in the cause of infection of M.grisea by complementary assays. All transformants restored the phenotype of wild type by one gene with 171bp away from the insertion site as well as the two genes.No sequence was found to be identical with the gene through nucleotide BLAST. The putative protein has 328aa with GENSAN analysis and has a 37~47% identities with a Zn-finger protein encoded by KIN17 from human, mouse, Dwsophila melanogaster and Arabidopsis. The sequence has a Zn-finger motif and a nuclear localization signal. 987bp ORF of the gene was obtained from the susceptible cDNA library. The research on gene knockout, the regulation of the gene expression and subcellular localization of the product is going on.