Xiaoru Fan

Plant nitrate transporters were first identified and functionally characterized more than 20 years ago. They are encoded at least by four gene families, NRT1 (NPF), NRT2, CLC, and SLAC1/SLAH. In this review, we overview the functions of the nitrate transporters in relation to their potential use as targets for improving crop nitrogen use efficiency. These functions include their roles in root architecture and nutrient acquisition; vacuole nitrate and protein storage; nutrient allocation from source to sink; sensing both abiotic and biotic stresses; the ionic balance of nitrate with potassium, chloride and cellular pH; and the circadian clock-regulated carbon and nitrogen balance. We provide and discuss some examples of the use of nitrate transporter genes and their regulators in improving plant growth and development, nitrogen use efficiency, and resistance to some abiotic stresses. We propose several strategies for effectively using nitrate transporters to achieve higher crop yields and nitrogen use efficiency by using gene transformation or genome editing or molecular marker-assisted breeding.

BACKGROUND AND AIMS: There is increased evidence that partial nitrate (NO3-) nutrition (PNN) improves growth of rice (Oryza sativa), although the crop prefers ammonium (NH4+) to NO3- nutrition. It is not known whether the response to NO3- supply is related to nitrogen (N) use efficiency (NUE) in rice cultivars. Methods Solution culture experiments were carried out to study the response of two rice cultivars, Nanguang (High-NUE) and Elio (Low-NUE), to partial NO3- supply in terms of dry weight, N accumulation, grain yield, NH4+ uptake and ammonium transporter expression [real-time polymerase chain reaction (PCR)]. KEY RESULTS: A ratio of 75/25 NH4+ -N/NO3- -N increased dry weight, N accumulation and grain yield of 'Nanguang' by 30, 36 and 21 %, respectively, but no effect was found in 'Elio' when compared with those of 100/0 NH4+ -N/NO3- -N. Uptake experiments with 15N-NH4+ showed that NO3- increased NH4+ uptake efficiency in 'Nanguang' by increasing Vmax (14 %), but there was no effect on Km. This indicated that partial replacement of NH4+ by NO3- could increase the number of the ammonium transporters but did not affect the affinity of the transporters for NH4+. Real-time PCR showed that expression of OsAMT1s in 'Nanguang' was improved by PNN, while that in 'Elio' did not change, which is in accordance with the differing responses of these two cultivars to PNN. Conclusions Increased NUE by PNN can be attributed to improved N uptake. The rice cultivar with a higher NUE has a more positive response to PNN than that with a low NUE, suggesting that there might be a relationship between PNN and NUE.

Summary The nitrate () transporter has been selected as an important gene maker in the process of environmental adoption in rice cultivars. In this work, we transferred another native Os NAR 2.1 promoter with driving Os NAR 2.1 gene into rice plants. The transgenic lines with exogenous pOsNAR 2.1:Os NAR 2.1 constructs showed enhanced Os NAR 2.1 expression level, compared with wild type ( WT ), and 15 N influx in roots increased 21%–32% in response to 0.2 m m and 2.5 m m and 1.25 m m 15 NH 4 15 NO 3 . Under these three N conditions, the biomass of the pOsNAR 2.1:Os NAR 2.1 transgenic lines increased 143%, 129% and 51%, and total N content increased 161%, 242% and 69%, respectively, compared to WT . Furthermore in field experiments we found the grain yield, agricultural nitrogen use efficiency ( ANUE ), and dry matter transfer of pOsNAR 2.1:Os NAR 2.1 plants increased by about 21%, 22% and 21%, compared to WT . We also compared the phenotypes of pOsNAR 2.1:Os NAR 2.1 and pOsNAR 2.1:Os NRT 2.1 transgenic lines in the field, found that postanthesis N uptake differed significantly between them, and in comparison with the WT . Postanthesis N uptake ( PANU ) increased approximately 39% and 85%, in the pOsNAR 2.1:Os NAR 2.1 and pOsNAR 2.1:Os NRT 2.1 transgenic lines, respectively, possibly because Os NRT 2.1 expression was less in the pOsNAR 2.1:Os NAR 2.1 lines than in the pOsNAR 2.1:Os NRT 2.1 lines during the late growth stage. These results show that rice NO 3 – uptake, yield and NUE were improved by increased OsNAR2.1 expression via its native promoter.

The NO3- transporter plays an important role in rice nitrogen acquisition and nitrogen-use efficiency. Our previous studies have shown that the high affinity systems for nitrate uptake in rice is mediated by a two-component NRT2/NAR2 transport system. In this study, transgenic plants were successfully developed by overexpression of OsNAR2.1 alone, OsNRT2.3a alone and co-overexpression of OsNAR2.1 and OsNRT2.3a. In our field experiments, transgenic lines expressing p35S:OsNAR2.1 or p35S:OsNAR2.1-p35S:OsNRT2.3a constructs exhibited increased grain yields of approximately 14.1% and 24.6% compared with wild-type (cv. Wuyunjing 7, WT) plants, and the agricultural nitrogen use efficiency increased by 15.8% and 28.6%, respectively. Compared with WT, the 15N influx in roots of p35S:OsNAR2.1 and p35S: OsNAR2.1-p35S:OsNRT2.3a lines increased 18.9% - 27.8% in response to 0.2 mM, 2.5 mM 15NO3– and 1.25 mM 15NH415NO3, while there was no significant difference between p35S:OsNAR2.1 and p35S:OsNAR2.1-p35S:OsNRT2.3a lines; only the 15N distribution ratio of shoot to root for p35S:OsNAR2.1-p35S:OsNRT2.3a lines increased significantly. However, there were no significant differences in nitrogen use efficiency, 15N influx in roots and the yield between the p35S:NRT2.3a transgenic lines and WT. This study indicated that co-overexpression of OsNAR2.1 and OsNRT2.3a could increase rice yield and nitrogen use efficiency.

Drought is an important environmental factor that severely restricts crop production. The high-affinity nitrate transporter partner protein OsNAR2.1 plays an essential role in nitrate absorption and translocation in rice. Our results suggest that OsNAR2.1 expression is markedly induced by water deficit. After drought stress conditions and irrigation, compared with wild-type (WT), the survival rate was significantly improved in OsNAR2.1 over-expression lines and decreased in OsNAR2.1 RNAi lines. The survival rate of Wuyunjing7 (WYJ), OsNRT2.1 over-expression lines and OsNRT2.3a over-expression lines was not significantly different. Compared with WT, overexpression of OsNAR2.1 could significantly increase nitrogen uptake in rice, and OsNAR2.1 RNAi could significantly reduce nitrogen uptake. Under drought conditions, the expression of OsNAC10, OsSNAC1, OsDREB2a, and OsAP37 was significantly reduced in OsNAR2.1 RNAi lines and increased substantially in OsNAR2.1 over-expression lines. Also, the chlorophyll content, relative water content, photosynthetic rate and water use efficiency were decreased considerably in OsNAR2.1 RNAi lines and increased significantly in OsNAR2.1 over-expression lines under drought conditions. Finally, compared to WT, grain yield increased by about 9.1% and 26.6%, in OsNAR2.1 over-expression lines under full and limited irrigation conditions, respectively. These results indicate that OsNAR2.1 regulates the response to drought stress in rice and increases drought tolerance.