[Effects of methyl jasmonate treatment on the hydrolytic activity and phosphorylation level of plasma membrane H+-ATPase in mung bean (Vigna radiata L.) hypocotyls].
Employing both protein kinase inhibitors and phosphatase inhibitors to investigate the effect of MeJA (methyl jasmonate) on the H+-ATPase hydrolytic activity of plasma membrane, and the regulations involved with phosphorylation and dephosphorylation of plasma membrane H+-ATPase after treatments with MeJA. 3 d-old etiolated mung bean (Vigna radiata L.) seedlings were harvested and the hypocotyls (1-2 cm in length) under the hook were used to prepare the plasma membrane vesicles by means of aqueous two-phase partition. Hydrolytic activities of plasma membrane H+-ATPase were determined in responding to the treatment with MeJA. H(+)-ATPase activity stimulated by MeJA and FC was up to 30%, and the combination of MeJA and FC did not show significant additive effect. The phosphatase inhibitors, okadaic acid and cantharidin, enhanced MeJA-induced increase of the enzyme activity to 60% and 50%, respectively. Staurosporine and cheleythrine, two inhibitors of protein kinase, abolished completely the stimulative effect of MeJA on PM H+-ATPase activity (Fig. 2). The results from gamma-(32)P tracer experiments showed that treatment with MeJA and FC increased the level of isotope labeling on PM H+-ATPase. Okadaic acid and cantharidin could enhance the labeling of gamma-(32)P on PM H+-ATPase induced by MeJA and FC. Both MeJA- and FC-induced increase in gamma-(32)P level were inhibited when cheleythrine was applied (Fig. 3). Ca2+ strongly stimulated the PM H+-ATPase and the increase of the enzyme activity was two times higher than that of the control. But Ca2+ had no enhancement of the enzyme activity induced by FC and MeJA. In the presence of Ca2+, Okadaic acid could increase the MeJA stimulation slightly, but cantharidin had not any significant effect on the enzyme activity. Staurosporine and cheleythrine showed no effect on MeJA-induced increase in PM H+-ATPase, and which activity was similar to the control after treatment with either inhibitor (Figs. 5, 6). The changes in hydrolytic activity of H+-ATPase stimulating by MeJA is probably related to reversible phosphorylation, and the protein kinase is Ca2+-independent, phosphatase is Ca2+-dependent.
Wen
Bin
B
Institute of Piwei, Guangzhou University of Traditional Chinese Medicine, Guangzhou 510405, China. wen_bin04@yahoo.com.cn
Bin
Jin-Hua
JH
Wang
Xiao-Jing
XJ
D000085:Acetates / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects* / Q000201:enzymology*; D003517:Cyclopentanes / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D007887:Fabaceae / Q000187:drug effects* / Q000201:enzymology*; D054883:Oxylipins / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D010937:Plant Growth Regulators / Q000494:pharmacology*; D006180:Proton-Translocating ATPases / Q000378:metabolism*
D000085:Acetates; D003517:Cyclopentanes; D054883:Oxylipins; D010937:Plant Growth Regulators; C072239:methyl jasmonate; D006180:Proton-Translocating ATPases; D002118:Calcium