The role of calcium in ABA-induced gene expression and stomatal movements.

There is much interest in the transduction pathways by which abscisic acid (ABA) regulates stomatal movements (ABA-turgor signalling) and by which this phytohormone regulates the pattern of gene expression in plant cells (ABA-nuclear signalling). A number of second messengers have been identified in both the ABA-turgor and ABA-nuclear signalling pathways. A major challenge is to understand the architecture of ABA-signalling pathways and to determine how the ABA signal is coupled to the appropriate response. We have investigated whether separate Ca2+-dependent and -independent ABA-signalling pathways are present in guard cells. Our data suggest that increases in [Ca2+]i are a common component of the guard cell ABA-turgor and ABA-nuclear signalling pathways. The effects of Ca2+ antagonists on ABA-induced stomatal closure and the ABA-responsive CDeT6-19 gene promoter suggest that Ca2+ is involved in both ABA-turgor signalling and ABA-nuclear signalling in guard cells. However, the sensitivity of these pathways to alterations in the external calcium concentration differ, suggesting that the ABA-nuclear and ABA-turgor signalling pathways are not completely convergent. Our data suggest that whilst Ca2+-independent signalling elements are present in the guard cell, they do not form a completely separate Ca2+-independent ABA-signalling pathway.

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 important component of Ca2+-based signal transduction pathways. This fact has led us to investigate whether oscillations in [Ca2+]cyt are involved in the response of stomatal guard cells to the plant hormone abscisic acid (ABA). We show that ABA induces oscillations in guard-cell [Ca2+]cyt. The pattern of the oscillations depended on the ABA concentration and correlated with the final stomatal aperture. We examined the mechanism by which ABA generates oscillations in guard-cell [Ca2+]cyt by using 1-(6-[17beta-3-methoxyestra-1,3, 5(10)-trien-17-yl]aminohexyl)-1H-pyrrole-2,5-dione (U-73122), an inhibitor of phosphoinositide-specific phospholipase C (PI-PLC)-dependent processes in animals. U-73122 inhibited the hydrolysis of phosphatidylinositol 4,5-bisphosphate by a recombinant PI-PLC, isolated from a guard-cell-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 both ABA-induced oscillations in [Ca2+]cyt and stomatal closure. In contrast, U-73122 did not inhibit external Ca2+-induced oscillations in guard-cell [Ca2+]cyt and stomatal closure. Furthermore, there was no effect of the inactive analogue 1-(6-[17beta-3-methoxyestra-1,3, 5(10)-trien-17-yl]aminohexyl)-2,5-pyrrolidinedione on recombinant PI-PLC activity or ABA-induced and external Ca2+-induced oscillations in [Ca2+]cyt and stomatal closure. This lack of effect suggests that the effects of U-73122 in guard cells are the result of inhibition of PI-PLC and not a consequence of nonspecific effects. Taken together, our data suggest a role for PI-PLC in the generation of ABA-induced oscillations in [Ca2+]cyt and point toward the involvement of oscillations in [Ca2+]cyt in the maintenance of stomatal aperture by ABA.