The Studies Of The Methyl Jasmonate-induced H₂O₂ Generation And Mediation In Guard Cells Of Arabidopsis Thaliana

Methyl-Jasmonate (MJ)is an endogenous signal for inducing stomatal closure. but until now we haven't seen any remark on the mechanism of this phenomenon from the point of view of signal transduction. we focus on the study of the MJ-induced H₂O₂ generation mediated by Ca²⁺ and MAPK guard cells of Arabidopsis thaliana by applying epidermis strip bioassay and laser scanning confocal microscopy. Epidermal bioassay demonstrated that 10-7-10-3mol/L MJ and H₂O₂ can induce stomatal closure in Arabidopsis thaliana guard cells. and 10^-5mol/L is the best effect on stomatal closure . CAT (100U/ml), Ascorbic acid (5mmol/L) and DPI (10μmol/L) all partially mimic the effect of MJ on stomatal closure after 3h treatment of 10^-5mol/L MJ. Confocal H₂O₂ mapping with the probe H2DCF-DA reveals that MJ (10^-5mol/L) leads to increasing in cytoplasmic in guard cells of Arabidopsis thaliana after 5 minutes, but the fluorescence of control has not change . And CAT, Ascorbic acid, and DPI can all partially reduce the production of MJ-induced H₂O₂. These results indicate that H₂O₂ can be involved in the regulation of MJ-induced stomatal closure . Further more, 10^-5mol/L MJ and 0.1mmol/L A₂₃₁₈₇ can make the fluorescence of DCF intense. After MJ (10^-5mol/L) leads to increasing in cytoplasmic H₂O₂, 0.5mmol/L EGTA and 1mmol/L LaCl3 can both make the fluorescence of DCF reduce. And 10^-5mol/L MJ and 10^-5mol/L H₂O₂ can make the fluorescence of Ca²⁺ in cytoplasmic in guard cells intense . After MJ (10^-5mol/L) leads to increasing in cytoplasmic Ca²⁺ , 100U/ml CAT,10^-5mol/L DPI and 5mmol/L Vc can all make the fluorescence of Ca²⁺ reduce. These data suggest that H₂O₂ and Ca²⁺ can be interdependent and interactive in MJ-regulated stomatal movement. MJ- or H₂O₂-induced stomatal closure was inhibited or reversed by the inhibitor PD98059 of MEK1/2 in Arabidopsis thaliana guard cells; After MJ-induced stomatal closure, the treatment of PD98059 could induce it open again, and MJ could not enhance the fluorescence intensity of H₂O₂ probe. These results suggest that MEK1/2 could mediate stomatal closure by the MJ-induced H₂O₂ generation/accumulation. In all, these data suggest that H₂O₂ as an intermediate can be involved in signal transduction pathways of MJ-regulated stomatal movement, and H₂O₂ and Ca2+ can be interdependent and interactive in MJ-regulated stomatal movement,and MAPK could mediate stomatal closure by the MJ-induced H₂O₂ generation/accumulation.