Functional Analysis Of Rice Blast Resistant Genes OsMP1 And OsREM1.2

Plants have a complex and diverse immune defense system to deal with pathogen infection,which involves the transmission of signals,energy and substances.Therefore,the study of disease-resistant signaling pathway related proteins can help reveal the mechanism of plant and pathogen interaction and provide a basis for the exploration of sustainable disease control.Rice blast is one of the main diseases of rice,and two rice blast resistance related proteins,OsMP1 and OsREM1.2,have been cloned in previous study.In this study,the functions of the OsMPl and OsREM1.2 were analyzed,and the main results were as follows:OsMP1 is an membrane protein gene that is obviously induced by rice blast inoculation identified from the microarray and iTRAQ proteomics in previous study.Subcellular localization experiment results show that OsMP1 is expressed on the cell membrane,and the bioinformatics predicted that OsMP1 containing 9 TMS(transmembrane a helix),and 2 repeat conservative EamA domain structure,belongs to the metabolism of drug/transporter superfamily(DMT),According to the analysis to the evolutionary tree between homologous genes,OsMP1 homologous genes mostly belongs to the MtN21 family.Coexpression analysis revealed that OsMP1 co-expressed with receptor kinases and NBS-LRR disease-resistant proteins,and was induced by blast fungus.In the ripening stage of rice,the plant height of OsMP1-overexpression transgenic lines is lower than that of the wild type,and the tiller number of transgenic lines based on CRISPR/Cas9 site-specific mutation is less than that of the wild type.When rice seedling was inoculated with blast fungus(Hoku 1),the average lesion length of OsMP1-overexpression transgenic lines is shorter than that of the wild type(Suyunuo),and the average lesion length of transgenic lines based on CRISPR/Cas9 site-specific mutation is longer than that of the wild type(Heikezijing).OsMP1::GFP fusion protein was aggregated at the site where the secondary mycelia invaded the adjacent cells after inoculation with blast fungus(Hoku 1).OsREM1.2 is a member of the remorin family,and is a kind of protein located in plasma membrane microregion(lipid raft).Six phosphorylation sites were predicted at the N end,with two peaking between 0.75-1.OsREM1.2 is homologous to the proteins(NbREM1.3,NtREM1.2 and StREM1.3)in tobacco and potato that inhibit viral infection in plants,and their amino acid sequences are conserved at the C end.Bioinformatics predicted that OsREM1.2 was highly expressed in internode,leaf,stem,root and callose of rice.In rice seedling inoculation,the average lesion length of OsREM1.2-overexpression transgenic lines is shorter than that of the wild type(Suyunuo),and the average lesion length of transgenic lines based on CRISPR/Cas9 site-specific mutation is longer than that of the wild type(Heikezijing).After inoculation with rice blast fungus(Hoku 1),the fusion expression protein of OREM1.2::GFP aggregate at the invasion point of appressorium,and the adjacent cells of secondary mycelium invasion.And It is expressed on the cell membrane of mycelium growth.In this study,the structure and function of two types of proteins related to plasma membrane were predicted,and the materials with overexpression and site-specific mutation based on CRISPR/Cas9 were constructed through transgenic means.After phenotype identification,they were found to be related to growth and development as well as resistance to rice blast.The expression protein of fused GFP was induced by rice blast,and expressed on the cell membrane,and aggregated at the invasion site.