Abscisic acid and hydrogen peroxide induce modification of plasma membrane H(+)-ATPase from Cucumis sativus L. roots under heat shock.

We examined the effect of heat shock (HS), for 2 h at 48°C, on plasma membrane H(+)-ATPase (PM-H(+)-ATPase) measured as the hydrolytic and H(+)-pumping activity. Some of the plants were transferred after 2 h HS to control temperature for another 24 h, as post-stressed (PS) plants. A significant increase of PM-H(+)-ATPase in plants subjected to HS was observed. The stimulation of PM-H(+)-ATPase was higher in PS plants. Estimation of transcript levels of cucumber PM-H(+)-ATPase in roots indicated that the action of HS affected gene expression levels. Transcript levels of two isoforms, CsHA4 and CsHA8, in PS plants were elevated. The expression of PM-H(+)-ATPase genes was not affected in plants treated for 2 h with HS. HS elevated the endogenous level of abscisic acid (ABA) both in plants treated for 2 h with HS and in PS plants. Moreover, in PS plants, a distinctly higher level of H(2)O(2) was observed. It was also demonstrated that transcript levels of PM-H(+)-ATPase were elevated in cucumber roots after 24-h treatment of plants with ABA or H(2)O(2). Both of these compounds seem to play an important role in increasing ATPase activity during heat stress, because the use of the inhibitors tungstate and DPI restrained stimulation of PM-H(+)-ATPase activity by heat. Moreover, protein blot analysis with an antibody against phosphothreonine and 14-3-3 protein indicated that increased activity of PM-H(+)-ATPase under HS resulted from phosphorylation of the enzyme. Taken together, the data presented here suggest that, under post-heat stress conditions, abscisic acid and hydrogen peroxide are involved in PM-ATPase modification, through stimulation of gene expression of that PM proton pump. Moreover, heat treatment of cucumber plants results in increased phosphorylation of PM-ATPase and thus fast post-translational modification, leading to activation of the enzyme protein.