Crosstalk of hydrogen sulfide and nitric oxide requires calcium to mitigate impaired photosynthesis under cadmium stress by activating defense mechanisms in Vigna radiata

Hydrogen sulfide (H₂S) and nitric oxide (NO) have been known to affect vast number of processes in plants under abiotic stresses. Also, calcium (Ca) works as a second messenger in plants, which underpins the abiotic stress-induced damage. However, the sequence of action of these signaling molecules against cadmium (Cd)-induced cellular oxidative damage remains unidentified. Therefore, we studied the synergistic actions and/or relationship of signaling molecules and Ca-dependent activation of tolerance mechanisms in Vigna radiata seedlings under Cd stress. The present study shows that exogenous Ca supplemented to Cd-stressed V. radiata seedlings reduced Cd accumulation and improved the activity of nitrate reductase, and L/D-cysteine desulfhydrase (LCD/DCD) that resulted in improved synthesis of NO and H₂S content. Application of Ca also elevated the level of cysteine (Cys) by upregulating the activity of Cys-synthesizing enzymes serine acetyltransferase and O-acetylserine(thiol)lyase in Cd-stressed seedlings. Maintenance of Cys pool under Cd stress contributed to improved H₂S content which together with Ca and NO improved antioxidant enzymes and components of ascorbate-glutathione (AsA-GSH) cycle. All these collectively regulated the activity of NADPH oxidase and glycolate oxidase, resulting in the inhibition of Cd-induced generation of reactive oxygen species. The elevated level of Cys also assisted the Cd-stressed seedlings in maintaining GSH pool which retained normal functioning of AsA-GSH cycle and led to enhanced content of phytochelatins coupled with reduced Cd content. The positive effect of these events manifested in an enhanced rate of photosynthesis, carbohydrate accumulation, and growth attributes of the plants. On the contrary, addition of NO scavenger cPTIO [2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide], H₂S scavenger HT (Hypotaurine) and Ca-chelator EGTA (Ethylene glycol-bis(b-aminoethylether)-N,N,N',N'-tetraacetic acid) again developed a condition similar to stress and positive effect of the signaling molecules was abolished. The findings of the study postulate that Ca in association with NO and H₂S mitigates Cd-induced impairment and enhances the tolerance of the V. radiata plants against Cd stress. The results of the study also substantiate that Ca acts both upstream as well as downstream of NO signals whereas, H₂S acts downstream of Ca and NO during Cd-stress responses of the plants.