Two potential Ca2+-dependent transduction pathways in stomatal closing in response to abscisic acid

Withering of abscisic acid (ABA)-insensitive mutants in standard watering conditions as well as under drought suggests that ABA would regulate stomatal closing not only under water stress but also under other hydric conditions. Depending on its applied concentration, the ABA closing signal might be transduced through different pathways. This possibility was investigated in epidermal peels of Commelina communis by comparing the effects of different Ca2+ buffers, protein kinase inhibitors, calmodulin (CaM) antagonists and guanosine-triphosphate-binding protein (G protein) modulators on the stomatal closing responses to 10 nM ABA (ABA10) and 100 nM ABA (ABA100). EGTA specifically suppressed the response to ABA10, while the Ca2+ buffer 1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA) similarly inhibited the responses to ABA10 and ABA100. The response to ABA10 was specifically affected by the protein kinase inhibitors KT5926 and 1-(5-iodonaphthalene-1-sulphonyl)-1H-hexahydro-1,4-diazepine (ML-7), the CaM antagonist N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), and the phospholipase C inhibitor 1-[6-[[17β-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-dione (U73122), whereas the response to ABA100 was specifically affected by the G protein antagonist pGlu-GlnD-Trp-Phe-D-Trp-D-Trp-Met-NH2 (GP Ant-2) and mas 17, an inactive mastoparan analog. These data are consistent with two possible ABA routes, each of them exhibiting specific Ca2+-requirements and protein phosphorylations, and being initiated by its own receptor. Furthermore, analogies with animal features suggest that the ABA10 and ABA100 closing signals might be extracellularly perceived.