Abscisic acid (ABA) is a well-studied regulator of stomatal movement. Hydrogen sulfide (H 2 S), a small signaling gas molecule involved in key physiological processes in mammals, has been recently reported as a new component of the ABA signaling network in stomatal guard cells. In Arabidopsis (Arabidopsis thaliana), H 2 S is enzymatically produced in the cytosol through the activity of L-cysteine desulfhydrase (DES1). In this work, we used DES1 knockout Arabidopsis mutant plants (des1) to study the participation of DES1 in the cross talk between H 2 S and nitric oxide (NO) in the ABA-dependent signaling network in guard cells. The results show that ABA did not close the stomata in isolated epidermal strips of des1 mutants, an effect that was restored by the application of exogenous H 2 S. Quantitative reverse transcription polymerase chain reaction analysis demonstrated that ABA induces DES1 expression in guard cell-enriched RNA extracts from wild-type Arabidopsis plants. Furthermore, stomata from isolated epidermal strips of Arabidopsis ABA receptor mutant pyrabactin-resistant1 (pyr1)/pyrabactin-like1 (pyl1)/pyl2/pyl4 close in response to exogenous H 2 S, suggesting that this gasotransmitter is acting downstream, although acting independently of the ABA receptor cannot be ruled out with this data. However, the Arabidopsis clade-A PROTEIN PHOSPHATASE2C mutant abscisic acid-insensitive1 (abi1-1) does not close the stomata when epidermal strips were treated with H 2 S, suggesting that H 2 S required a functional ABI1. Further studies to unravel the cross talk between H 2 S and NO indicate that (1) H 2 S promotes NO production, (2) DES1 is required for ABA-dependent NO production, and (3) NO is downstream of H 2 S in ABA-induced stomatal closure. Altogether, data indicate that DES1 is a unique component of ABA signaling in guard cells.Abscisic acid (ABA) regulates diverse physiological and developmental processes in plants, among which seed dormancy and stomatal movement are the most studied. Stomata are pores bordered by pairs of specialized cells named guard cells located in the epidermis of the aerial part of most land plants. Due to the waxy cuticle of plants, stomatal pores regulate approximately 90% of all the gas exchange (i.e. the uptake of CO 2 required for photosynthesis and the loss of water vapor during transpiration) between the plant and the environment (Hetherington and Woodward, 2003). Thus, stomatal movement is a key process for the regulation of plant water status and biomass production. In guard cells, ABA induces an increase of intracellular calcium concentrations, which, in turn, induces an efflux of anions that causes membrane depolarization. In this voltage milieu, ion uptake is blocked through the inactivation of inward-rectifying potassium channels, and ion efflux is induced through activation of outward-rectifying potassium channels. This solute relocation drives water out of the guard cells and closes the stomatal pore as a result of a reduction in guard cell turgor (Blatt, 2000;Kim et al...