The Function And Mechanism Of Soybean Resistance Gene GmRpsYu Against Soybean Phytophthora Root Rot

The soybean Phytophthora root rot caused by Phytophthora sojae is a worldwide soil-borne disease that harms soybean production and seriously affects soybean yield and quality.Planting resistant soybean varieties is the most economical,effective and environmentally friendly method to control soybean Phytophthora root rot.Therefore,it is very critical to screen soybean Phytophthora root rot resistant germplasm resources and discover and utilize resistant genes.In the early stage of this study,a hybrid population of Phytophthora root rot resistant Yudou 29 and susceptible Jikedou 2 was constructed,and the candidate resistant gene GmRpsYu was obtained by map-based cloning and sequencing.Structural analysis revealed that GmRpsYu encodes a C₂H₂-type zinc finger protein transcription factor.Through functional verification and mechanism analysis,it was proved that GmRpsYu mediated soybean resistance to soybean Phytophthora root rot through peroxide pathway.The results of the study are as follows:1.Analysis of expression profile showed that GmRpsYu was mainly expressed in stems and leaves of soybean,and the expression in leaves of Yudou 29 was significantly higher than that of Jikedou 2.GmRpsYu expression was clearly induced by the infection of P.sojae.In addition,treatment of Yudou 29 leaves with pathogen-associated molecular patterns（PAMPs）flg22 and chitin induced expression of GmRpsYu.GmRpsYu positively regulates soybean resistance to P.sojae.After inoculation with P.sojae,GmRpsYu overexpression plants with genetic background of susceptible varieties showed strong resistance.Therefore,GmRpsYu was identified as the resistance gene of soybean Phytophthora root rot.GmRpsYu was localized in the nucleus of plant cells by subcellular localization.2.GmRpsYu also showed resistance to bacterial blight of soybean caused by Pseudomonas syringae pv.glycinea.The peroxidase pathway is involved in soybean resistance to bacterial blight of soybean:POD and SOD activities in Yudou 29 were significantly higher than in Jikedou 2 after inoculation,the SOD activity was significantly higher in the GmRpsYu overexpression plants than in Williams 82.The expression levels of three SOD1 of the four cultivars were significantly increased after inoculation.The results showed that SOD1 gene was involved in soybean disease resistance pathway.3.GmRpsYu is involved in PAMPs-induced soybean immune response.Using flg22and chitin treatment of Williams 82 and GmRpsYu overexpression plants that GmRpsYu positively regulated the expression of immune response related genes PR1-1,JAR4,NAC6and ICS1.4.RNA-Seq analysis was performed on Williams 82 and GmRpsYu overexpression plants inoculated with P.sojae and uninoculated.Under different conditions,the genes whose log2（Fold Change）|>1&qvalue<0.05 are regarded as differentially expressed genes（DEGs）.2578 up-regulated DEGs and 972 down-regulated DEGs were found after Williams82 inoculation;1779 up-regulated DEGs and 432 down-regulated DEGs were found after GmRpsYu overexpression plant inoculation.GmRpsYu overexpression plants uninoculated treatment had 29 up-regulated DEGs and 61 down-regulated DEGs compared with Williams82.GmRpsYu overexpression plants inoculated with P.sojae had 29 up-regulated DEGs and83 down-regulated DEGs compared with Williams 82.DEGs are mainly involved in transcription,metabolic and biosynthesis of secondary metabolites pathways by GO function and KEGG analysis.Given that RNA-seq showed that the peroxide pathway may be involved in the resistance to soybean Phytophthora root rot,the expression of SOD genes was verified by q RT-PCR with similar trends of RNA-Seq results.5.Resistance of GmRpsYu overexpression plants to P.sojae mediated by peroxidase pathway:The POD and SOD activity of Yudou 29 inoculated with P.sojae were significantly higher than that of Jikedou 2.SOD activity and H₂O₂content of GmRpsYu overexpression plants were significantly higher than that of Williams 82.Exogenous SOD substantially enhanced P.sojae-resistance of Jikedou 2 and Williams 82.Yeast one-hybrid,GUS staining and luciferase experiments demonstrated that GmRpsYu as a transcription factor could bind to and activate SOD1-03 and SOD1-19 promoters.