| Potato is the fourth most important food crop in the world.Late blight is a devastating disease of potatoes,which seriously threatens the development of the potato industry.So how to improve potato late blight resistance is a difficult and urgent task in the world potato breeding program.It is very important to explore late blight resistance genes of potato and investigate their molecular mechanism.Previous studies showed that the receptor-like kinase St LRPK1,with leucine-rich repeat sequence,interacts with the co-receptor St BAK1 to positively regulate late blight resistance in potato and Nicotiana benthamiana.However,St LRPK1 may not possess kinase activity due to the mutated key sites in kinase structural domain.Therefore,we speculated that St LRPK1 would interact with other receptor kinases,and then they form a complex with co-receptor St BAK1 to conduct its function.Aimed to find its’ interaction components,IP-MS was used to find candidate interacting protein of St LRPK1,and a L-type plant lectin receptor-like kinase StLecRK.Ⅸ was found.Based on the above results,this study investigated the late blight resistance function of StLecRK.Ⅸ,and further investigated the mechanism of StLecRK.Ⅸ/St LRPK1/St BAK1 form a protein complex to synergistically regulating the potato late blight resistance.The main findings of this study are as follows:1.Evolutionary tree and structural domain analysis showed that the homologs of StLecRK.Ⅸ from Arabidopsis,N.benthamiana and tomato share conserved structural domains,they all contain an extracellular lectin domain and an intracellular kinase domain.Further comparison revealed that StLecRK.Ⅸ also contains an N-terminal signal peptide and a transmembrane domain.Subcellular localization observation confirmed that StLecRK.Ⅸ was localized on the cell membrane.2.StLecRK.Ⅸ expression profile was examined in potato.It was abundantly expressed in potato roots and up-regulated in leaves in response to the induction of CF,the culture filtrate of P.infestans PAMPs.Kinase activity assays of prokaryotic expressed protein confirmed that StLecRK.Ⅸ possess kinase activity.3.The interactions between StLecRK.Ⅸ and St LRPK1,StLecRK.Ⅸ and St BAK1,and St LRPK1 and St BAK1 were confirmed by Co-immunoprecipitation and luciferase complementation assay.The results indicated that StLecRK.Ⅸ/St LRPK1/St BAK1 may function in plants by forming a ternary protein complex and CF treatment did not affect their interaction.4.Stable overexpression of StLecRK.Ⅸ in potato significantly increased P.infestans resistance,but interference of StLecRK.Ⅸ had no effect on that.Both transient overexpression of StLecRK.Ⅸ and stable overexpression of Nb Lec RK.Ⅸ in N.benthamiana significantly improved resistance against P.infestans,on the contrary,the resistance was significantly decreased in Nb Lec RK.Ⅸ interfered plants.5.StLecRK.Ⅸ overexpression stimulates basal immune response in potato and N.benthamiana,including reactive oxygen species burst,callose deposition and expression of defense related genes such as St PTI5,St WRKY8 and St HSR3.In summary,this study demonstrated that potato StLecRK.Ⅸ,St LRPK1 and St BAK1 positively regulate late blight resistance in potato and N.benthamiana by forming a ternary protein complex,and StLecRK.Ⅸ could stimulate immune responses,such as reactive oxygen species burst,callose deposition,and expression of defense related genes.This study enriched the research information of receptor-like kinases in regulating potato late blight resistance and provided a basis for using StLecRK.Ⅸ/St LRPK1 to improve potato late blight resistance. |
