Cloning And Biological Function Of G Protein Signaling Factor CgRGS1、CgRGS2 And CgRGS7 In Colletotrichum Gloeosporioides

Rubber farming industry has a very important position in China’s hot agricultural production, They were not only the basic industries of Hainan economy and the advantages of industry, but also the majority of diligent farmers to increase the pillar industries.However, colletotrichum caused by rubber anthracnose is one of the major diseases of rubber trees, to the growth of the rubber tree caused serious harm and will reduce the annual production of rubber very Seriously. And lead to the disease of the main pathogenic fungi - Colletotrichum is not only a wide range of growth,but also have variety of ways to infect host. Thus,in-depth study of the bacteria in the pathogenesis of related genes in the expression and regulation of the network, to understand the interaction mechanism between Colletotrichum gloeosporioides and host, and provide effective theoretical guidance for effective control of the pathogen.Regulators of G-protein signaling (RGS) are a kind of negative regulatory factor of G protein, which play important roles in growth development and pathogenicity of plant pathogen. RGS domain is a key for RGS protein to play a role in GTPase activation. In the previous study of the laboratory, we had identified 10 potential RGS proteins from Colletotrichum gloeosporioides,They all contain the RGS functional domain,three RGS genes of Colletotrichum gloeosporioides were cloned on the basis of this study, Through the analysis of its protein sequence, and the corresponding biological traits, to further confirm its biological function. The results obtained are as follows:1. Three genes were obtained by cloning, and they were found to be highly homologous to MoRGS1,MoRGS2 and MoRGS7 genes in Magnaporthe oryzae. So they were named CgRGS1,CgRGS2 and CgRGS7. The gene-knockout mutant of CgGS1、CgRGS2 and CgRGS7 was obtained by homologous recombination.2.The gene of CgRGS1:Bioinformatics analysis results show that the length of the ORF of CgRGSl is 2208 bp and encoded a 735-amino acids protein, there are contains two DEP functional domains in the N terminus and containing a RGS function domain in the C terminal. Alignment analysis showed that amino acid sequence of CgRGS1 is 99%similar with that of CgRGS1 from C. gloeosporioides Nara gc5. Comparing to the wild type, the knockout mutant of CgRGSl had slow growth, conidium production, formation of appressorium and the reaction is more sensitive in the hypertonic environment. The protein CgRGS1 was involved in regulation of vegetative growth, conidium production and germination, cell wall integrity and pathogenicity of C. Gloeosporioides.3.The gene of CgRGS2:Bioinformatics analysis results show that the length of theORF of CgRGS2 is 1725 bp and encoded a 574-amino acids protein, containing a RGS function domain in the N terminal. Alignment analysis showed that amino acid sequence of CgRGS2 is 91% similar with that of CgRGS2 from C. gloeosporioides Nara gc5.Comparing to the wild type, the knockout mutant of CgRGS2 had slow growth, thick aerial hyphae, reduced conidia with multi-end germination, sensitive to oxidative stress and SDS, decreased pathogenicity. The protein CgRGS2 was involved in regulation of vegetative growth, conidium production and germination, oxidative stress response, cell wall integrity and pathogenicity of C. Gloeosporioides.4.The gene of CgRGS7:Bioinformatics analysis results show that the length of the ORF of CgRGS7 is 1860 bp and encoded a 620-amino acids protein, There are 7 transmembrane domains in the N terminus and containing a RGS function domain in the C terminal. Comparing to the wild type, the knockout mutant of CgRGS7 reduced conidia with multi-end germination, decreased appressorium formation rate and pathogenicity of C. Gloeosporioides.The above results indicated that CgRGS7 gene involved in the regulation of conidial production,formation of appressorium and pathogenicity of Colletotrichum gloeosporioides.