| Kiwifruit canker is caused by Pseudomonas syringaepv.actinidiae(PSA)with 6different biovars;however,among thesebiovars,biovar 3 isconsidered as major.There arevery few studies conducted on the functions of the effectors secreted by PSA T3 SS.Thus,current study focused on the functions of the effectors secreted by the PSA biovar3.The screened effectors bioinformatics analyses,andthe effectors expression quantity in the process of infection,and heterologous expression and interaction proteins were carried out.In addition,the specific effector Hop Z5 was used to design specific primers and construct a quantitative PSA detection system to improve the detection sensitivity.Based on the above analyses,the results so far achieved are summarized as follows:1.Six effectors of PSA biovar3,such as Hop H1,Hop AM1-2,Hop Z5,Hop Z3,Hop BB1-1,Hop BB1-2,were observed for the preliminary functional prediction.Hop H1 belongs to the Peptidase M19 family and is a zinc-dependent endopeptidase;while Hop AM1-2 has no conserved domain.Both Hop Z5 and Hop Z3 belong to the Yersinia effector Yop J family,inhibited innate immune of plants,block the MAP kinase and NFκB signaling pathways by acting on the Phosphorylation site.The functional domains of Hop BB1-1 and Hop BB1-2 were similar to the functional domain of Avr Pph F,contain ORF2,which is toxic to plants.The expression levels of all were the highest after 3 days of inoculation of PSA on Hongyang.It shows that they played a certain role in the process of infection.2.The six effecters were distinctly different from other pathogenic species,including the specific effector of Hop Z5.Subcellular localization revealed that Hop H1,Hop AM1-2,Hop Z5,Hop Z3 and Hop BB1-2 were localized to the plasma membranes and nucleus;while Hop BB1-1 only observed in plasma membranes.The leaves of N.benthamiana showed cell death when expression of Hop Z5 occurs after 48 h;while Hop H1 showed after 72 h,and the remaining effectors after 96 h.In order to study the interaction between effectors and plant,the effectors Hop Z3 and Hop BB1-2 were expressed in N.benthamiana,and it was found that the interacting proteins of the effectors were mainly participated in secondary metabolite biosynthesis,oxidative phosphorylation pathway,and so on.3.Hop Z5 is the specific effector of PSA biovar3 and for the specificity of the effector,a real-time fluorescence quantitative detection system was constructed,which could improve the sensitivity by 100 times compared with the normal PCR.The system was applied to detect PSA in branches without obvious symptoms in different time periods under different environmental conditions,and the PSA in the greenhouse were reproducing prior to it in natural conditions. |
