Abscisic acid stimulation of phospholipase D in the barley aleurone is G-protein mediated and localized to the plasma membrane.

Sian Ritchie and Simon Gilroy

Biology Department, 208 Mueller Laboratory, The Pennsylvania State University, University Park, Pennsylvania 16802

Abstract.
We have previously determined that phospholipase D (PLD) is activated by abscisic acid (ABA) and this activation is required for the ABA response of the cereal aleurone cell. In this study, ABA stimulated PLD activity was reconstituted in vitro in microsomal membranes prepared from aleurone protoplasts. The transient nature (20 min) and degree (1.5-2 fold) of activation in vitro were similar to that measured in vivo. Stimulation by ABA was only apparent in the membrane fraction, and was associated with a fraction enriched in plasma membrane. These results suggest that an ABA receptor system, and elements linking it to PLD activation, is associated with the aleurone plasma membrane. The activation of PLD in vitro by ABA was dependent on the presence of GTP. Addition of GTPgS transiently stimulated PLD in an ABA independent manner, whereas treatment with GDPbS or pertussis toxin (PTX) blocked the PLD activation by ABA. Application of PTX to intact aleurone protoplasts inhibited the ability of ABA to activate PLD, as well as antagonizing the ability of ABA to down-regulate gibberellic acid-stimulated a-amylase production. All these data support the hypothesis that ABA stimulation of PLD activity occurs at the plasma membrane and is mediated by G-protein activity.