Study On The Pathogenicity-related Functional Domain VRpep Of Chinese Wheat Mosaic Virus And Compatative Analysis Of SiRNA Derived From Wheat Yellow Mosaic Virus In Different Tissues Of Wheat

Chinese wheat mosaic virus(CWMV)and Wheat yellow mosaic virus(WYMV)are two important wheat viruses transmitted by Polymyxa graminis,which are responsible for wheat yellow mosaic disease and cause great yield loss in winter wheat growing region of China.Nearly all the protein encoded by plant virus might function as pathogenicity determinant,and viral RNA replicase contribute to the pathogenesis during plant virus infection has been reported for several plant viruses.Recent studies indicated that virus derived small interfering RNA(vsiRNA)can also cleavage target endogenous mRNA of host,and responsible for the symptom development of the host plant.In this study,a pathogenicity-related functional domain named VRpep,but not vsiRNA,is identified from variable region of replicase in CWMV genome which can enhance pathogenesis for various plant viruses in Nicotiana benthamiana.Preliminary study was performed on the pathogenesis mechanism of VRpep.In addition,analysis of vsiRNA from WYMV infected roots and leaves in wheat plant demonstrated that RNA silencing in roots may operate differently than in leaves against soil-borne virus invasion.The results of this study are listed as following:1.A pathogenicity-related functional domain named CR1-7 is identified from replicase P153 of CWMV through screening of pathogenicity-related functional domain in CWMV replicase region and pathogenicity vsiRNA in the whole CWMV genome.Experiments of deletion mutation and early termination of translation suggests that the function of CR1-7 is because of the functional domain rather than vsiRNA sequences.Meanwhile,we shortened the length of the effective functional domain to 46 amino acid with the name VRpep.2.Furthermore,besides Tobacco rattle virus(TRY),CWMV VRpep can also promote infection and replication of Potato virus X(PVX)and Tobacco mosaic virus(TMV),increasing the accumulation level of viral mRNA and protein in N.benthamiana.This result suggests that the pathogenicity enhancement by VRpep might not specific to one plant virus.3.Multiple alignment analysis indicated that CWMV VRpep and homologous sequences of viruses in Furovirus are highly conservative.In addition,VRpep homologous sequences of Furovirus can also effectively promote infection and replication of TRV in N.benthamiana,suggesting the function of VRpep might be Furovirus specific.Moreover,three conserved continually amino acid of VRpep homologous sequences(FLD)might play essential roles for the virulence.4.Both CWMV VRpep and CR1-7 are not silencing suppressor,and cannot increase silencing suppressor activity of other viruses(TRV,PVX)as well,indicating that pathogenic enhancement mechanism of VRpep(CR1-7)might not related to RNAi pathway of host plant.5.88 proteins in N.benthamiana were identified to be potentially interacted with CWMV CR1-7 through Pull-down experiment.GO analysis revealed that several proteins are involved in defense pathway of host plant,suggesting that CR1-7(or VRpep)might interact with corresponding protein and inhibit host defense related pathway.6.Characteristics analysis of WYMV siRNAs derived from roots and leaves of wheat plant indicated that there are obvious differences between the two tissues for the vsiRNA abundance,distribution pattern along the viral genome and 5’-terminal nucleotide.In addition,RNAi related genes(DCL like genes)were differently regulated between infected leaves and roots,suggesting that RNA silencing in roots may operate differently than in leaves against soil-borne virus infection in wheat plant.WYMV and CWMV are two important wheat viruses in China,results in this study will contribute to the future studies on the interaction between soil-borne viruses and host plants,and provide new potential strategies for the control of soil-borne wheat disease in China.