Functional Analysis Of Guanine Kinase And Impdh Active Sites In Magnaporthe Oryzae

Rice blast disease caused by Magnaporthe oryzae,is one of the three major diseases,and it has become a threaten to the safety of the global rice,therefore the control of rice blast is always an important subject in the production of rice.The main prevention and control of rice blast is breeding resistant varieties,supplemented by chemical control.Because the pathogen races are unstable,resistant varieties will be lost after planted in the field for a period of time.Therefore,the prevention and control of rice blast is one of the difficult problems in rice production.To understand the molecular mechanism of Magnaporthe oryzae growth and pathogenic process is an important prerequisite for the control.With the publication of the complete genome sequence of Magnaporthe oryzae,using molecular biology methods to research function of pathogenicity related gene is becoming more and more convenient,which provides the basis for finding new targets for fungicides.Guanosine triphosphate(GTP)plays a very important role in vivo.First of all,GTP is a crucial material for the synthesis of RNA,and also involved in the synthesis of cyclic guanosine monophosphate(cGMP)which plays the intracellular second messenger role in vivo.Additionally,during the process of GTP synthesis pathway can provide energy for vesicle transport and signal transduction etc.Here,we characterized functions of MoGukl and MoImd4 which play important roles in GTP biosynthesis pathway in M.oryzae.In the pathway of the synthesis of GTP,guanylate kinase can be considered as a housekeeper.In the first chapter,we mainly aimed at biological function of the guanylate kinase of Magnaporthe oryzae,MoGuk1.We obtained ?Mogokl mutant by gene knockout,compared with the wild type strain Guyll,the mutant showed a reduced growth rate in vegetative growth,the asexual development decreased significantly and loss the ability of sexual reproductive,meanwhile the germination of the spores germination and appresorium formation rate decreased,and the turgor pressure became lower.Besides the infection hyphae in the host cells,expansion capacity and pathogenicity of ?Moguk1 mutant was significantly reduced.The results showed that,MoGuk1 play an important role in growth,asexual and sexual development and pathogenicity in M.oryzae.There are two purine synthesis pathway in vivo,one is de novo synthesis pathway,the other is salvage pathway,and the main way is de novo synthesis pathway.Inosine-5'-monophosphate dehydrogenase(IMPDH)is a key enzyme in GTP de novo synthesis pathway and it is a rate-limiting reaction in relation to the later steps.IMPDH catalytic dehydrogenation and hydrolysis reaction of GTP de novo synthesis pathway,and have different conformations in different catalytic reactions.The second chapter of this paper mainly aimed at the inosine-5'-phosphate dehydrogenase MoImd4,of which we carried out point mutations on ten avtive sites and analysis their biological functions,we found that the pathogenicity of point mutations ?Moimd4/MoImD4T268AR269A??Moimd4/MoIMD4D290A??Moimd4/MoIMD4Y428A??Moimd4/MoImD4M431AG432A??Moimd4/MoIMDQ470A are the sare as ?Moimd4.This indicates that these sites play very important role in the catalytic process of IMPDH.