Effects of in Vivo Treatment with Abscisic Acid and/or Cytokinin on Activities of Vacuolar H<sup>+</sup> Pumps of Tonoplast-Enriched Membrane Vesicles Prepared from Barley Roots

Abstract

We investigated the effects of in vivo treatment (1 day) of barley roots with abscisic acid (ABA) and/or a cytokinin (6-benzyladenine; BA) on the ATP- and PPi-dependent H+ transport activities of tonoplast-enriched membrane vesicles prepared from the roots. Treatment with ABA significantly increased the two H+ transport activities. By contrast, treatment with BA significantly decreased PPi-dependent H+ transport activity, while the change in ATP-dependent H+ transport activity was small. Increases in the two H+ transport activities caused by treatment with ABA were suppressed during treatment with ABA and BA. Changes in the NO−-inhibitable ATPase activity and the Na+-inhibitable PPiase activity of membrane vesicles after treatment of roots with phytohormone(s) (ABA, BA, ABA + BA) were similar to changes in the ATP- and PPi dependent H+ transport activities of the membrane vesicles, respectively. Immunoblot analysis with antibodies raised against the functional catalytic subunits of the vacuolar H+ pumps (H+- ATPase and H+-PPiase) of mung bean revealed that only the level of the functional catalytic subunit of the H+-PPiase of the membrane vesicles was significantly increased by treatment with ABA alone and in combination with BA. These results suggest that treatment with ABA has a stimulatory effect on the activities of the two H+ pumps of the vacuolar membrane of barley roots, with increase in the level of the catalytic subunit of the H+-PPiase, and that treatment with BA has an inhibitory effect on the two H+ pump activities of the vacuolar membrane without changes in the levels of the catalytic subunits of either H+ pump, with the limitation that treatment with BA has an inhibitory effect only when the activity of the H+-ATPase has been increased by treatment with ABA.