Melatonin-mediated endogenous nitric oxide coordinately boosts stability through proline and nitrogen metabolism, antioxidant capacity, and Na+/K+ transporters in tomato under NaCl stress

Abstract

The interactions between nitric oxide (NO) and melatonin in alleviating sodium chloride (NaCl) toxicity in plants are poorly comprehended. Here, the associations between the exogenous application of melatonin and endogenous NO levels in inducing tomato seedlings' defense response during NaCl toxicity were investigated. The results indicated that the application of melatonin (150 μM) increased height (23.7%) and biomass (32.2%), improved chlorophyll (a (137%) and b (92.8%)), and proline metabolisms, and reduced the contents of superoxide anion radicals (49.6%), hydrogen peroxide (31.4%), malondialdehyde (38%), and electrolyte leakage (32.6%) in 40-day-old tomato seedlings grown under NaCl (150 mM) treatment. Melatonin increased the antioxidant defense system in NaCl-stressed seedlings by increasing the activity of the antioxidant enzymes. Melatonin also improved N metabolism and endogenous NO content in NaCl-stressed seedlings by upregulating the activity of enzymes implicated in N assimilation. Furthermore, melatonin improved ionic balance and reduced Na content in NaCl-exposed seedlings by upregulating the expression of genes involved in K/Na ratio homeostasis (NHX1-4) and increasing the accumulation of mineral nutrients (P, N, Ca, and Mg). However, the addition of cPTIO (100 μM; an NO scavenger) reversed the beneficial impacts of melatonin, indicating the effective function of NO in melatonin-induced defense mechanisms in NaCl-stressed tomato seedlings. Therefore, our results revealed that melatonin improves the tolerance of tomato plants during NaCl toxicity by mediating internal NO.

Keywords: NHX genes; NaCl stress; Solanum lycopersicum; melatonin; nitric oxide; nitrogen metabolism.

Figure 1

The leaf contents of minolevulinic acid (ALA, A) and proline (B), and the leaf activities of aminolevulinic acid dehydratase (ALAD, C), chlorophyllase (Chlase, D), pyrroline-5-carboxylate synthetase (P5CS, E), and proline dehydrogenase (ProDH, (F) enzymes in tomato seedlings treated with melatonin (150 μM) and cPTIO (100 μM) under NaCl (150 mM) treatment. Different letters in each column show significant differences at the p < 0.05 based on the LSD test (means ± SD, n = 5), CT, control treatment; M, melatonin; S, salinity.

Figure 2

The leaf contents of hydrogen peroxide (H₂O₂, A), superoxide anion radical (B), malondialdehyde (MDA, C), and electrolyte leakage (EL, D) in tomato seedlings treated with melatonin (150 μM) and cPTIO (100 μM) under NaCl (150 mM) treatment. Different letters in each column show significant differences at the p < 0.05 based on the LSD test (means ± SD, n = 5), CT, control treatment; M, melatonin; S, salinity.

Figure 3

The leaf activities of catalase (CAT, A), superoxide dismutase (SOD, A), ascorbate peroxidase (APX, B), and glutathione reductase (GR, B) enzymes in tomato seedlings treated with melatonin (150 μM) and cPTIO (100 μM) under NaCl (150 mM) treatment. Different letters in each column show significant differences at the p < 0.05 based on the LSD test (means ± SD, n = 5), CT, control treatment; M, melatonin; S, salinity.

Figure 4

The leaf content of nitric oxide (NO, A) and the leaf activities of nitrate reductase (NR, B), nitrite reductase (NiR, B), glutamine synthetase (GS, C), and glutamyl synthetase (GOGAT, D) enzymes in tomato seedlings treated with melatonin (150 μM) and cPTIO (100 μM) under NaCl (150 mM) treatment. Different letters in each column show significant differences at the p < 0.05 based on the LSD test (means ± SD, n = 5), CT, control treatment; M, melatonin; S, salinity.

Figure 5

The relative expression of NHX1 and NHX2 genes in the root (A, C) and leaf (B, D) of tomato seedlings treated with melatonin (150 μM) and cPTIO (100 μM) under NaCl (150 mM) treatment. Different letters in each column show significant differences at the p < 0.05 based on the LSD test (means ± SD, n = 3), CT, control treatment; M, melatonin; S, salinity.

Figure 6

The relative expression of NHX3 and NHX4 genes in the root (A, C) and leaf (B, D) of tomato seedlings treated with melatonin (150 μM) and cPTIO (100 μM) under NaCl (150 mM) treatment. Different letters in each column show significant differences at the p < 0.05 based on the LSD test (means ± SD, n = 3), CT, control treatment; M, melatonin; S, salinity.