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Huai-Fu Fan, Chang-Xia Du and Shi-Rong Guo

Nitric oxide (NO), an endogenous signaling molecule in plants and animals, mediates responses to abiotic and biotic stresses. This study was conducted in a nutrient solution to investigate 1) the effects of exogenous sodium nitroprusside (SNP), an NO donor, on free proline (Pro) and protein content; and 2) the enzymes involved in Pro metabolism [pyrroline-5-carboxylate synthetase (P5CS) and proline dehydrogenase (PDH)] in cucumber (Cucumis sativus) seedling leaves and roots under NaCl stress. The results showed that the increases in free Pro and protein were significantly higher in the 50 mm NaCl solution but highly significant with the addition of 100 μM SNP to the 50 mm NaCl solution for the entire treatment period. Moreover, leaves maintained higher levels of free Pro and protein content than roots throughout the experiments. The P5CS activity increased in the saline treatment compared with the control, and this increase was greater in the 50 mm NaCl + 100 μM SNP solution than in the other treatments. On the other hand, the PDH activity was inhibited under NaCl stress but the reduction in activity was greater in the 50 mm NaCl + 100 μM SNP solution than in the others. These findings suggest that Pro metabolism was significantly altered during the exogenously applied NO under salt stress and that this alteration prompted the accumulation of higher levels of free Pro, which, in turn, maintained the turgor in the cucumber seedlings and helped protect them from salt stress. Moreover, the toxic effects generated by 50 mm NaCl were partially overcome by the application of NO, which could be used as a potential growth regulator to improve plant salinity tolerance. Therefore, it was concluded that NO could alleviate salinity damage in cucumber seedlings by regulating Pro metabolism. Overall, the adverse effects of salt stress could be lessened by the exogenous application of NO to cucumber seedlings.

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Huai-Fu Fan, Wen Chen, Zhou Yu and Chang-Xia Du

Salt stress reduces the fresh weight, dry weight, and relative growth rate of cucumber (Cucumis sativus) seedlings and results in serious quality loss in cucumber production. Our previous study indicated that the netting-associated peroxidase (CsaNAPOD) protein in cucumber seedling roots was induced by salt stress. Here, we amplified the coding sequence of CsaNAPOD from a cDNA isolated from the roots of cucumber seedlings. Sequence analysis indicated that the coding sequence of CsaNAPOD is 1035 bp, encoding a deduced protein of 344 amino acids, with a predicated molecular weight of 37.2 kD and theoretical isoelectric point of 5.64. The deduced amino acid sequence of CsaNAPOD showed high sequence similarity to peroxidases (PODs) from other plant species. Moreover, CsaNAPOD possesses the typical sequence structures of class III PODs and indicated that CsaNAPOD belongs to this subfamily. CsaNAPOD was highly expressed in the roots and was weakly expressed in the stems and leaves of cucumber seedlings. Salt stress significantly increased the expression of CsaNAPOD in the leaves during the entire experimental period compared with the control, and the expression of CsaNAPOD in roots was reduced at 6 hours and induced at 48 and 72 hours by salt treatment. In stems, the expression of CsaNAPOD declined at 48 and 72 hours as a result of the salt treatment compared with the control. These results indicate that the expression of CsaNAPOD responded to salt stress in cucumber seedlings, and the expression patterns under salt stress in different tissues were not identical. Our research suggests that CsaNAPOD may have potential function during the plant response to salt stress.

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Chang-Xia Du, Huai-Fu Fan, Shi-Rong Guo and Takafumi Tezuka

To examine whether spermidine (SPD) modifies plant antioxidant enzyme expression in response to short-term salt stress, cucumber (Cucumis sativus) seedlings were treated with NaCl in the presence or absence of SPD for 3 days. Compared with untreated control plants, free radical production and malondialdehyde content in leaves and roots increased significantly and plant growth was suppressed under 50 mm NaCl stress. Exogenous SPD sprayed on leaves at a concentration of 1 mm alleviated salinity-mediated growth reduction. Salt stress caused a consistent increase in soluble protein content, as well as peroxidase (POD) and superoxide dismutase (SOD) activities in cucumber seedlings. By native polyacrylamide gel electrophoresis, five POD isozymes were detected in cucumber seedling leaves, and seven in roots. We detected five SOD isozymes in leaves and four in roots, and two catalase (CAT) isozymes in leaves and two in roots. Our results indicate that salt stress induced the expression of POD and SOD isozymes in cucumber seedlings, but inhibited the expression of CAT isozymes in roots. Application of exogenous SPD further increased POD and SOD expression and activity, and led to the differential regulation of CAT in leaves and roots. These data show that antioxidant enzymes, especially POD and SOD, appear to protect cucumber seedlings against stress-related damage, and they appear to function as the molecular mechanisms underlying the response of cucumber seedlings to salinity. Moreover, SPD has potential to scavenge directly free radical and to alleviate growth inhibition and promote the activity and expression of antioxidant system enzymes in cucumber seedlings under short-term salt stress.