Nitrate reductase-dependent nitric oxide production mediates nitrate-conferred salt tolerance in rice seedlings

Soil salinity is a destructive environmental factor that inhibits plant growth and crop yield. Applying nitrogen fertilizer is a practical method to enhance salt tolerance. However, the underlying mechanisms remain largely unknown. Here, we demonstrated that NO3--enhanced salt tolerance in rice (Oryza sativa L.) seedlings is mediated by nitrate reductase (NR)-dependent nitric oxide (NO) production. Seedlings grown in nitrate condition (N) exhibited much greater salt tolerance compared with those grown in ammonium nitrate and ammonium (A) conditions, a pattern also observed in the MADS-box transcription factor 27 (mads27) mutant. NR activity was highly induced by NO3- under both normal and salt stress conditions. Only the double mutant nr1/2 and the triple mutant nr1/2/3 displayed a dramatic reduction in salt tolerance. Application of tungstate suppressed salt tolerance of wild-type seedlings but not the triple mutants. Furthermore, both NO3--enhanced salt tolerance and salt-induced NO production were totally blocked in triple mutants. However, treatment with exogenous sodium nitroprusside (an NO donor) significantly enhanced salt tolerance in both Nipponbare (NIP) and the triple mutants. Antioxidant enzyme activities in shoots were significantly inhibited in the triple mutants when compared with NIP. Furthermore, expression of OsAKT1 was specifically induced by NO3- but was inhibited in the roots of triple mutants, resulting in a lower potassium/sodium ratio in NR triple mutants. Our results revealed that NO3--conferred salt tolerance is mediated by NR-dependent NO production in rice seedlings. Nitrate application promotes nitrate reductase activity and enhances nitric oxide production, which activates antioxidant enzymes in rice shoots, increases the K+/Na+ ratio in rice seedlings, and confers salt tolerance.