Study On The Action Mechanism Of PcF/SCR Effector SCR96 Involving In The Pathogenesis Of Phytophthora Cactorum

PCF/SCR(small cystine-rich proteins)is one group of extracellular effector proteins of Phytophthora pathogens,which can cause plant cell death(PCD).However,the action mechanismof these proteins is still unclear.Our previous studies demonstrated that the PCF/SCR effector SCR96 is a virulence factor of P.cactorum.In this thesis,the following studies were carried out to further analyze the action mechanism of SCR96 in the interaction between P.cactorum and plants,so as to provide an important basis for elucidating the pathogenesis mechanism of P.cactorum.In this thesis,SCR96 tagged with His was expressed in Pichiapastoris and purified using Ni-NTA.The protein activity was analyzed by infiltrating tomato leaves with the recombinant SCR96(530,265,133 and 66.5 nM).The results showed that after optimizing the protein expression system,the recombinant SCR96 at a concentration of 150 ?M was obtained;tomato PCD induced by SCR96 was concentration-dependent,and no PCD could be observed when SCR96 was of less than 66.5 nMPlant immune responses(reactive oxygen species burst,callose deposition,activation of defense-related genes and inoculation reaction)were determined when tomato leaves were treated with the recombinant SCR96.The results showed that the recombinant SCR96 could induce reactive oxygen species burst,callose deposition and defense-related gene expression changes in tomato leaves.Plant resistance against Botrytis cinerea and P.capsici was promoted by exogenously spraying SCR96.The effect of Agrobacterium-mediated transient expression of SCR96 on the expression levels of plant intercellular proteins was investigated using Label-free proteomics.The results showed that the expression levels of 36 proteins located in the plant apoplast and plasma membrane were significantly changed.Plant hydrogen peroxide catalytic process and peroxidase activity pathway were influenced by SCR96 treatment.These results indicate that SCR96 is related to reactive oxygen species pathway in plants.The plant interaction proteins of SCR96 were identified by yeast two-hybrid screening,and verified by bimolecular fluorescence complementation(BiFC).Using Agrobacterium-mediated expression system,plants transiently expressing interaction proteins were inoculated with pathogens to analyze their functions involved in the disease resistance.The results showed that SCR96 probably interacts with a suberization-associated anionic peroxidase-like POD.Its overexpression promoted the susceptibility of Nicotiana benthamiana to P.capsici infection.This thesis confirmed that the PCF/SCR effector SCR96 can induce plant immunity,which leads to significant expression changes of proteins related to resistance in plant extracellular space;moreover,SCR96 compromises plant disease resistance by targeting plant anionic peroxidase.The results provide important data for elucidating the action mechanism of SCR96 in the pathogenesis,and lay a solid foundation for further exploration of the interaction mechanisms between Phytophthora pathogens and plants.