Hydrogen sulfide generated by L-cysteine desulfhydrase 1 enhances plant basal thermotolerance through the regulation of stomatal behavior and the promotion of photosynthetic efficiency

Heat stress (HS) severely limits plant growth and crop yields, becoming a critical challenge for global food production. As a gasotransmitter, hydrogen sulfide (H₂S) has been reported to participate in plants HS adaptation, however, the underlying mechanism remains unclear. Time-course analysis showed that HS duration-dependently boosted endogenous H₂S production, with both transcriptional upregulation and enzymatic activation of the L-Cysteine desulfhydrase 1 (DES1). Over-expression of DES1 enhanced basal thermotolerance in Arabidopsis, whereas the DES1 knockout mutant (des1) exhibited heightened sensitivity to HS. Additionally, HS triggered stomatal closure and elevated stomatal density were substantially attenuated in des1, but more pronounced in OE-DES1. Importantly, the heat-sensitive phenotype and defective HS-induced stomatal responses in des1 could be rescued by exogenous H₂S pretreatment. Under HS condition, H₂S generated by DES1 triggered stomatal closure by regulating the expression of K⁺_in channel encoding genes, at least KAT1 and KC1, thereby modulating K⁺ homeostasis and turgor pressure of guard cells. Concurrently, DES1-H₂S up-regulated the expression of genes associated stomatal development, SPCH and TMM, increasing stomatal density. Correspondingly, des1 had decreased level of relative water content, chlorophyll levels, and RuBisCO activity during HS, while OE-DES1 had the opposite effects. Overall, H₂S might act as an equilibrator of stomatal aperture and stomatal density to avoid excessive transpiration and maximize gaseous exchange then photosynthetic efficiency of leaves under HS. Our study elucidates how H₂S optimizes stomatal behavior to improve basal thermotolerance, providing new insights into H₂S signaling and potential applications in breeding heat-resistant crops.