Study On The Function And Regulatory Mode Of G-protein β Subunit MGB1 In Maize

Plant G protein consists of three subunits:Gα,Gβ and Gγ,which regulate multiple biological processes by interacting with downstream effector proteins.Maize contains one typical Ga protein,three atypical extra-large Gα(XLG)proteins,one Gβ protein and six Gγ proteins.In this experiment,the function and regulation mode of MGB1 were studied.The main results are as follows:1.Sequence characterization and functional analysis of maize G protein βsubunit MGB1:MGB1 contains seven trypto-phan aspartic acid 40(WD40)repeat domains.The N-terminal of MGB1 has a coiled-coilhelices domain,which is necessary for the formation of Gβγ dimer with Gγ subunits.The prediction of the tertiary structure of MGB1 protein showed that the repetitive motif of WD40 formed a β propeller structure.The results of yeast two-hybrid showed that MGB1 interacted with MGG4.MGB1 was constitutively expressed in roots,leaves and grains.We used CRISPR/Cas9 technology to obtain editing materials KO1 and KO2 of MGB1.Wild-type plants WT,MGB1 editing materials KO1 and KO2 could germinate normally.KO1 and KO2 died at seedling stage,indicating that MGB1 was necessary for maize seedling establishment.2.Study on transcriptional regulation of maize G protein β subunit MGB1:in this experiment,wild type plant WT and MGB1 editing material KO1 were selected for transcriptome sequencing,and the transcriptional regulation network of MGB1 was analyzed.The results showed that compared with WT,a total of 5192 differentially expressed genes were found in transgenic material KO1,including 2957 up-regulated genes and 2235 down-regulated genes.GO analysis and KEGG analysis of differentially expressed genes showed that differentially expressed genes were significantly enriched in defense response pathway.We further analyzed 9 PR proteins(pathogenesis-related protein,)and 3 RPM proteins(plant immune receptor)by qRT-PCR.The results of RNA-seq and qRT-PCR were consistent,which showed that compared with WT,the expression of 9 PR genes and 3 RPM genes increased in KO1.3.Interaction model analysis of maize G protein β subunit MGB1:protein interaction network analysis showed that MGB1 might interact with MAPK pathway elements MAPK1,MAPK3 and MAPKK2.The results of yeast two-hybrid split-LUC showed that MGB1 interacted with RACK1,RACK1 interacted with MAPK elements MAPK1 and MAPK3,but not with MAPKK2.This study revealed that MGB1 is related to plant defense response at the transcriptome level.The construction of MGB1 transcriptional regulatory network lays a foundation for further understanding the regulation mechanism of maize G protein and provides a new perspective for the analysis of the interaction of G protein-MAPK pathway.