Reactive carbonyl species function as signal mediators downstream of H₂O₂ production and regulate [Ca²⁺]_cyt elevation in ABA signal pathway in Arabidopsis guard cells

We have demonstrated that reactive carbonyl species (RCS) function as an intermediate downstream of hydrogen peroxide (H₂O₂) production in abscisic acid (ABA) signaling for stomatal closure in guard cells using transgenic tobacco plants overexpressing alkenal reductase. We investigated the conversion of the RCS production into downstream signaling events in the guard cells. Both ABA and H₂O₂ induced production of the RCS, such as acrolein and 4-hydroxy-(E)-2-nonenal (HNE), in epidermal tissues of wild-type Arabidopsis thaliana plants. Application of the RCS scavengers, carnosine and pyridoxamine, did not affect the ABA-induced H₂O₂ production but inhibited the ABA- and H₂O₂-induced stomatal closure. Both acrolein and HNE induced stomatal closure in a plasma membrane NAD(P)H oxidase mutant atrbohD atrbohF as well as in the wild type, but not in a calcium-dependent kinase mutant cpk6. Acrolein activated plasma membrane Ca²⁺-permeable cation channels, triggered cytosolic free Ca²⁺ concentration ([Ca²⁺]_cyt) elevation, and induced stomatal closure accompanied by depletion of glutathione in the guard cells. These results suggest that RCS production is a signaling event between the ROS production and [Ca²⁺]_cyt elevation during guard cell ABA signaling.