The phytohormone abscisic acid (ABA) plays pivotal roles in triggering stomatal closure and facilitating adaptation of plants to drought stress.Hydrogen sulfide (H2S),a small signaling gas molecule,is involved in ABA-dependent stomatal closure.However,how H2S regulates ABA signaling remains largely unclear.Here,we show that ABA induces the production of H2S catalyzed by L-CYSTEINE DESULFHYDRASE1 (DES1) in guard cells,and H2S in tu positively regulates ABA signaling through persulfidation of Open Stomata 1 (OST1)/SNF1-RELATED PROTEIN KINASE2.6 (SnRK2.6).Two cysteine (Cys) sites,Cys131 and Cys137,which are exposed on the surface of SnRK2.6 and close to the activation loop,were identified to be persulfidated,which promotes the activity of SnRK2.6 and its interaction with ABA response element-binding factor 2 (ABF2),a transcription factor acting downstream of ABA signaling.When Cys131,Cys137,or both residues in SnRK2.6 were substituted with serine (S),H2S-induced SnRK2.6 activity and SnRK2.6-ABF2 interaction were partially (SnRK2.6c131S and SnRK2.6C137S) or completely (SnRK2.6c131SC137S) compromised.Introduction of SnRK2.6c131s,SnRK2.6c137s,or SnRK2.6c131SC137S into the ost1-3 mutant could not rescue the mutant phenotype: less sensitivity to ABA-and H2S-induced stomatal closure and Ca2+ influx as well as increased water loss and decreased drought tolerance.Taken together,our study reveals a novel post-translational regulatory mechanism of ABA signaling whereby H2S persulfidates SnRK2.6 to promote ABA signaling and ABA-induced stomatal closure.