Involvement of hydrogen sulfide (H2S) in stomatal immunity

SCUFFI, DENISE; GOTOR, CECILIA; LAXALT, ANA M.; AROCA, ANGELES; SCHWARZLÄNDER, MARKUS; PANTALENO, ROSARIO; COSTA, ALEX; GARCÍA-MATA, CARLOS

Mendoza

Congreso; LVIII Annual Meeting of the Argentine Society for Biochemistry and Molecular Biology Research; 2022

SAIB

Stomata are pores in the aerial part of the plants surrounded by a pair of cells, guard cells. The regulation of the pore size is crucial since it allows gas exchange with the environment and the loss of water by evapotranspiration. The regulation of stomatal aperture is key to maintain carbon and water homeostasis and also for plant defense since pathogens can enter the plant through the stomatal pore.Hydrogen sulfide (H2S) is a gaseous signaling molecule. In plants, the main enzymatic source is L-cysteine desulfhydrase 1 (DES1) which degrades L-cysteine to H2S, pyruvate and ammonia in the cytosol. The involvement of H2S has been reported both in response to stress and in development processes in plants. Today it is considered that H2S mechanism of action is given mainly by protein persulfidation, a redox posttranslational modification (PTM) in protein cysteine residues. This modification can modify the activity or localization of the target protein.In our lab, we have demonstrated that exogenous application of H2S induces stomatal closure in several plant species involving the participation of other signaling molecules such as nitric oxide (NO) and reactive oxygen species (ROS). Moreover, in Arabidopsis thaliana endogenous H2S/DES1 participates in pathways that lead to stomatal closure as abscisic acid (ABA) and the bacterial elicitor flagelin (flg22). Our principal aim is to unveil the signaling pathway and the mechanism of action of endogenously produced H2S that lead to stomatal closure in response to pathogen perception. Stomatal closure assays, with different pathogen elicitors in isolated epidermal peels of wild type and des1 mutant Arabidopsis leaves, show that des1 does not respond to flg22 but does respond to elf18 and chitin suggesting a specific role of DES1 in flg22 signaling. Moreover, we used the ratiometric Orp1-GFP2 biosensor and observed that flg22-induced H2O2 increase is blocked in presence of H2S scavenger, hypotaurine (HT). We further explored the cytosolic H2O2 dynamics in response to H2S in rbohD mutant and observed a response that was similar to that of wild type suggesting that H2O2 induction by H2S is RBOHD-independent. We also analyze apoplastic ROS burst in response to flg22 in leaf discs by luminol-based assay. The result demonstrates that DES1 participates in this response since des1 mutant induces significantly less ROS than wild type. Finally, we are performing a Dimedone-switch assay on Arabidopsis epidermal peels treated with flg22 in order to identify persulfidation targets. Preliminary results show several targets associated to the immune response, suggesting this PTM is an active mechanism in H2S–dependent stomatal closure in response to flg22.Further investigations are needed in order to know the H2S targets and the role of persulfidation during stomatal immunity.