Abstract
Stomatal opening is largely promoted by light-activated plasma membrane-localized proton ATPases (PM H+-ATPases), while their closure is mainly modulated by abscisic acid (ABA) signaling during drought stress. It is unknown whether PM H+-ATPases participate in ABA-induced stomatal closure. We established that BRI1-ASSOCIATED RECEPTOR KINASE 1 (BAK1) interacts with, phosphorylates and activates the major PM Arabidopsis H+-ATPase isoform 2 (AHA2). Detached leaves from aha2-6 single mutant Arabidopsis thaliana plants lost as much water as bak1-4 single and aha2-6 bak1-4 double mutants, with all three mutants losing more water than the wild-type (Columbia-0 [Col-0]). In agreement with these observations, aha2-6, bak1-4, and aha2-6 bak1-4 mutants were less sensitive to ABA-induced stomatal closure than Col-0, whereas the aha2-6 mutation did not affect ABA-inhibited stomatal opening under light conditions. ABA-activated BAK1 phosphorylated AHA2 at Ser-944 in its C-terminus and activated AHA2, leading to rapid H+ efflux, cytoplasmic alkalinization, and reactive oxygen species (ROS) accumulation, to initiate ABA signal transduction and stomatal closure. The phosphorylation-mimicking mutation AHA2S944D driven by its own promoter could largely compensate for the defective phenotypes of water loss, cytoplasmic alkalinization, and ROS accumulation in both aha2-6 and bak1-4 mutants. Our results uncover a crucial role of AHA2 in cytoplasmic alkalinization and ABA-induced stomatal closure during the plant's response to drought stress.