Extracellular ATP mediates H2S-regulated stomatal movements and guard cell K+ current in a H2O2-dependent manner in Arabidopsis

Hydrogen sulfide (H2S) is a newly discovered gaseous signaling molecule and involved in ethylene and ABA-induced stomatal closure. As an important factor, extracellular ATP (eATP) was believed to participate in regulation of stomatal closing. However, the mechanism by which eATP mediates H2S-regulated stomatal closure remains unclear. Here, we employed Arabidopsis wild-type and mutant lines of ATP-binding cassette transporters (Atmrp4, Atmrp5 and their double mutant Atmrp4/5) to study the function of eATP in H2S-regulated stomatal movement. Our results indicated that H2S affected stomatal closing through stimulating guard cell outward K+ current. Moreover, we found that H2S induced eATP generation by regulating the activity of an ABC transporter. The inhibitor of ABC transporters, glibenclamide (Gli), could impair H2S-regulated stomatal closure and reduce H2S-dependent eATP accumulation in Atmrp4 and Atmrp5 mutants. In addition, the promotion effect of H2S on outward K+ currents was diminished in Atmrp4/5 double mutant. Our data suggested that hydrogen peroxide (H2O2) is required for H2S-induced stomatal closure, and the production of H2O2 is regulated by eATP via NADPH oxidase. Based on this work, we conclude that H2S-induced stomatal closure requires ABC transporter-dependent eATP production and subsequent NADPH oxidase-dependent H2O2 accumulation.