Mechanisms Of NO And H₂O₂ In The Resistance Responses Of Cotton Suspension Cells To Toxin Of Verticilium Dahliae

In this study, an attempt is made to account for the observed difference in the susceptibility of Gossypium hirsutum L. (cv. BD-18 and Simian 3, resistant and susceptible to V. dahliae, respectively) to toxin of Verticillium dahliae, by looking for differences in the timing and level of production of NO and H₂O₂, and their effects on the expression of GST gene.In this experiment, a new method was used to purify the toxin protein produced by Verticillium dahliae. The supernatant of fungus culture was frozen and dried with Lyophilizer first, and then dialysed by Dialysis Membranes (MWCO 1000) after dissolved in distilled water. This method can eliminate the salt and sucrose in culture medium and reserve the protein component almost completely. VD-toxin were analysed using sodium dodecyl sulfate polyacrylamide gel analysis(SDS-PAGE). The results indicated that the protein components were very complex, and included glycoprotein within 35.8kDa-83.2kDa. Furthermore, treatments on glycoproteins with high temperature, conA and proteinase alleviated, not abolished, their activities.NO and H₂O₂ production were assayed in two cultivars of cotton cells which were treated with VD-toxin. The resistant cultivar exhibited higher induced levels of NO and H₂O₂ compared to the susceptible cultivar. The dual effects of NO were verified in this research. NO was either oxidative or antioxidative which depends on its concentration. NO took part in antioxidative system within appropriate concentration.GSTs are important enzymes in plants response to stress. We use SqRT- PCR to analyze the expression changes of GST gene in cotton suspension cultures. The result revealed that VD-toxin can induce the expression of GST gene, and the resistant cultivar exhibited higher and earlier induced levels compared to the susceptible cultivar. NO, H₂O₂ also induced the expression of GST gene, however NO induced defense responses against VD-toxin may independent on H₂O₂ signaling pathways leading to GST gene expression.