NO accumulation alleviates H2O2-dependent oxidative damage induced by Ca(NO3)2 stress in the leaves of pumpkin-grafted cucumber seedlings

Nitric oxide (NO) and hydrogen peroxide (H2O2), two important signaling molecules, are stimulated in plants by abiotic stresses. In this study, we investigated the role of NO and its interplay with H2O2 in the response of self-grafted (S-G) and salt-tolerant pumpkin-grafted (Cucurbita maxima x C. moschata) cucumber seedlings to 80 mM Ca(NO3)(2) stress. Endogenous NO and H2O2 production in S-G seedlings increased in a time-dependent manner, reaching maximum levels after 24 h of Ca(NO3)(2) stress. In contrast, a transient increase in NO production, accompanied by H2O2 accumulation, was observed at 2 h in rootstock-grafted plants. N-w-Nitro-L-Arg methyl ester hydrochloride (L-NAME), an inhibitor of nitric oxide synthase (NOS), tungstate, an inhibitor of nitrate reductase (NR), and 2-(4-carboxyphenyl)-4,4,5,5-tetramethy-limidazoline-1-oxyl-3-oxide (cPTIO), a scavenger of NO, were found to significantly inhibit NO accumulation induced by salt stress in rootstock- grafted seedlings. H2O2 production was unaffected by these stress conditions. Ca(NO3)(2) stress-induced NO accumulation was blocked by pretreatment with an H2O2 scavenger (dimethylthiourea, DMTU) and an inhibitor of NADPH oxidase (diphenyleneiodonium, DPI). In addition, maximum quantum yield of PSII (Fv/Fm), as well as the activities and transcript levels of antioxidant enzymes, were significantly decreased by salt stress in rootstock grafted seedlings after pretreatment with these above inhibitors; antioxidant enzyme transcript levels and activities were higher in rootstock- grafted seedlings compared with S-G seedlings. These results suggest that rootstock grafting could alleviate the oxidative damage induced by Ca(NO3)(2) stress in cucumber seedlings, an effect that may be attributable to the involvement of NO in H2O2-dependent antioxidative metabolism.