Abscisic acid induces oscillations in guard-cell cytosolic free calcium that involve phosphoinositide-specific phospholipase C

Oscillations in cytosolic free Ca2+ concentration ([Ca2+](cyt)) are an impo rtant component of Ca2+-based signal transduction pathways. This fact has l ed us to investigate whether oscillations in [Ca2+](cyt) are involved in th e response of stomatal guard cells to the plant hormone abscisic acid (ABA) . We show that ABA induces oscillations in guard-cell [Ca2+](cyt). The patt ern of the oscillations depended on the ABA concentration and correlated wi th the final stomatal aperture. We examined the mechanism by which ABA gene rates oscillations in guard-cell [Ca2+](cyt) by using 1-(6-{[17 beta-3-meth oxyestra-1,3,5(10) -trien-17-yl] amino}-hexyl)-1H-pyrrole-2,5-dione (U-7312 2), an inhibitor of phosphoinositide-specific phospholipase C (PI-PLC)-depe ndent processes in animals. U-73122 inhibited the hydrolysis of phosphatidy linositol 4,5-bisphosphate by a recombinant PI-PLC, isolated from a guard-c ell-enriched cDNA Library, in a dose-dependent manner. This result confirms that U-73122 is an inhibitor of plant PI-PLC activity. U-73122 inhibited b oth ABA-induced oscillations in [Ca2+](cyt) and stomatal closure. In contra st, U-73122 did not inhibit external Ca2+-induced oscillations in guard-cel l [Ca2+](cyt) and stomatal closure. Furthermore, there was no effect of the inactive analogue 1-(6- {[17 beta-3-methoxyestra-1,3,5 (10)-trien-17-yl] a mino} hexyl)-2,5-pyrrolidinedione on recombinant PI-PLC activity or ABA-ind uced and external Ca(2+-)induced oscillations in [Ca2+](cyt) and stomatal c losure. This lack of effect suggests that the effects of U-73122 in guard c ells are the result of inhibition of PI-PLC and not a consequence of nonspe cific effects. Taken together, our data suggest a role for PI-PLC in the ge neration of ABA-induced oscillations in [Ca2+](cyt) and point toward the in volvement of oscillations in [Ca2+](cyt) in the maintenance of stomatal ape rture by ABA.