Induced Resistance And Mechanism Of Protein Elicitor PevD1from Verticillium Dahliae In Cotton

PevD1, a protein elicitor, was isolated and purified from the culture medium of Verticillium dahliae.Previous research has indicated that the recombinant protein-PevD1, expressed in Escherichia coli,could activate the hypersensitive response （HR） and systemic acquired resistance （SAR） in tobaccoplants. In order to investigate the roles and mechanisms of PevD1elicitor in cotton resistance against V.dahlia, we used cotton and V.dahliae as research system in this work, measured the defense enzymesactivity and the relative content of secondary metabolites, and detected the expression ofpathogenesis-related （PR） genes and the key genes from the phenylpropanoid metabolism pathway incotton induced by PevD1with RT-PCR technique. Our results not only provide a supplemental methodfor controlling cotton Verticillium wilt disease, also provide a theoretical foundation for plant-elicitorinteractions and for elucidatation of the mechanisms of PevD1inducing plant resistance.1. PevD1induced SAR in cotton plants and improved plant resistance to V. dahliae. The recombinantprotein elicitor PevD1that expressed in E. coli reduced disease index of protein-treated plants,both resistant cultivar and susceptible variety. PevD1could not cause plant wilt and not aphytotoxic factor in tested cotton strain.2. Optimal inducing method, duration and concentration were determined. Different inducingmethods, duration and PevD1concentration were significantly affected the disease resistanceinduced by PevD1. The best induction condition was3days after treatment of PevD1leaf injectionat a concentration as low as8μg/mL.3. Elicitor PevD1activated defense response in cotton plant. PevD1systemically induced earlysignaling molecules production （hydrogen peroxide accumulation, nitric oxide generation） anddefensive substance deposition, such as lignin and terpenoids that function as fungal infectionsuppression and enhanced cotton resistance. PevD1could also induce the vessel reinforcement andproduce many granular materials filled in the contact cells.4. Defense enzyme activity in cotton treated with PevD1and inoculated with V. dahlia was compared.The results showed that the kinetic trend of phenylalanine ammonia-lyase （PAL）, polyphenoloxidase （PPO） and peroxidase （POD） activity in plant treated by PevD1was similar as treated by V.dahlia. The maximum activity of PAL appeared at96h and the activity of PPO and POD reachedthe peak at120h after PevD1treatment. The PAL and POD activity of V. dahlia-treated leavesreached a peak at120h. Similarly, PPO activity reached a peak at144h after treatment. The peaksof PPO and PAL activity in plant treated with V. dahlia were delayed than that with PevD1.5. Transcriptional level of PR genes in cotton treated with PevD1and inoculated with V. dahlia wasanalyzed with RT-PCR technique. The results indicated that PevD1stimulation and V. dahliainoculation could increase the transcriptional level of β^-1,3-glucanase, basic chitinase, acidicchitinase and cadinene synthase, and also increased the transcriptional level of key genes, such asPAL、C4H1and4CL from the phenylpropanoid metabolism pathway that mediated cotton defenseresponse. These results revealed that protein elicitor PevD1induced SAR and its functional mechanism, andalso demonstrated that PevD1act as an effector in cotton-pathogen V. dahlia interactions and trigger theplant immune responses. PevD1may act as an importment role in cotton-pathogen interactions.