Multiple roles of nitric oxide in root development and nitrogen uptake

ABSTRACT

Nitric oxide (NO) is widely recognized for its role as a signaling molecule in regulating plant developmental processes. We summarize recent work on NO generation via nitrate reductase (NR) or/and NO synthase (NOS) pathway in response to nutrient fluctuation and its regulation of plant root growth and N metabolism. The promotion or inhibition of root development most likely depends on NO concentrations and/or experimental conditions. NO plays an important role in regulating plant NR activity at posttranslational level probably via a direct interaction mechanism, thus contributing largely to N assimilation. NO also regulates N distribution and uptake in many plant species. In rice cultivar, NR-generated NO plays a pivotal role in improving N uptake capacity by increasing root growth and inorganic N uptake, representing a potential strategy for rice adaption to a fluctuating nitrate supply.

Pathways of NO synthesis in plants

Nitric oxide (NO) synthesis pathways can be classified as oxidative or reductive in operation.¹ The oxidative route involves mainly L-arginine-, polyamine- and hydroxylamine-mediated pathways. NO synthase (NOS) has been shown to be a main enzymatic source of NO production in animals.^2-4 A plant NOS has not been identified to date,^5-10 although experiments using inhibitors of animal NOS have provided evidence for a role of the L-arginine pathway in NO production in plants.¹¹ Reductive pathways also have three routes: the cytosolic nitrate reductase (NR), plasma membrane-bound nitrite:NO reductase and mitochondrial nitrite reduction pathways.¹ There is no doubt that NR is one of the most important sources of NO in plants.^12,13

Several lines of evidence suggest that enzymatic pathways (NOS and/or NR), are involved in NO production in response to nutrient fluctuations.^14-18 NO levels are likely enhanced via the NOS pathway in roots grown under phosphorus (P) and iron (Fe) deficiency.^14-18 Despite research showing that NOS rather than NR was involved in LN-induced NO production in maize,¹⁹ however, NO was shown to be generated through a NIA2-dependent NR pathway initially under N deficiency in rice. The main reason is that, at day 1 of N deficiency, nitrate accumulating in roots rather than being transported from the roots to the shoots led to elevate NR in root.¹⁴ And it's no doubt that, with N deficiency continuing, the main resource of enhanced NO should be shifted from NR to NOS pathway. Besides nutrient stress, nutrient supplies with nitrogen forms also can affect NO generation. The nitrate supply caused a consistent increase in NO fluorescence in the first minutes after treatment,^20-22 whereas there was little change due to ammonium supply throughout the 6-h experiment.⁹ Similarly, in rice plants which prefer ammonium over nitrate as a N source, NO levels are still enhanced in roots via a NIA2-dependent NR pathway when ammonium is partially substituted for by nitrate (partial nitrate nutrition, PNN).¹⁰

The role of NO in root growth

Nitric oxide (NO) as a signaling molecule involves in regulating plant root growth.^10,19,23-26 The promotion or inhibition of root development most likely depends on NO concentrations and/or the experimental conditions. For example, for primary root elongation, NO inhibitory effect is mostly derived from experiments of the exogenous application of NO donors^9,24-27 and NO-overproducing mutants;²⁸ NO promote effect mainly results from the experiments under stress experiments^{9,13,19,29-31} and mutant plants exhibiting lower NO levels.^32-34

Increasing studies have shown that NO participates root growth and development in response to nutrient fluctuation in plants.^11-14 A burst of NO triggered by magnesium deficiency stimulated the development of root hairs in Arabidopsis.²⁵ Higher levels of endogenous NO in P-deficient roots have been positively correlated with the development of cluster roots in white lupin.¹² NO as a shared signaling molecule in the formation of white lupin cluster roots induced by P and Fe deficiencies.¹³ Similarly, NO has been shown to be involved in N- and P-deficiency-induced seminal root elongation in rice plants via induction of meristem cell activity.¹⁴ It has been more than 50 years since nitrate was first shown to stimulate plant root growth,³⁵ it is easy to speculate that NR-generated NO participates nitrate-mediated root growth.^10,36

NO-induced N uptake and metabolism

Nitrate acts both as a nutrient and signal that regulates plant N acquisition and metabolism.^37-39 However, nitrate appears to have no direct effects on the modulation of NR activity but through the alteration of endogenous NO level.^20-22 NR activity in wheat leaf is negatively modulated by NO released from NO donor (SNP or GSNO); simultaneously nitrate content significantly increased, indicating that the substrate for NR activity was present in amounts enough to be not a limiting factor for NR activity.²² Further experiments showed that the haem and molybdenum centers in NR protein were the two sites modulated by NO,²¹ suggesting that the regulation of NR activity by NO is regulated at the post-translational level, probably via a direct interaction mechanism. Above results suggested that NO could play an important role in regulating NR activity in plants, thus contributing largely to N assimilation. Nevertheless, whether the effect of NO on NR activity is positive or negative largely dependent on experimental conditions. ^8,21-22 Increasing the rate of NO gas increased NR activity in the enzyme extracts of the roots fed with 0.5mM nitrate but decreased it when 5 mM nitrate was supplied, illustrating the complex regulation mechanism of NO on N metabolism.⁸

Besides NO regulation of NR activity, NO also regulates N uptake and distribution.^10,36,40 Recent evidence indicate a significant effect of NO on organic- and inorganic-N uptake and allocation of N to different N pools in the fine roots of beech seedlings, depending strongly on NO concentration, N availability and N source.^36,40-41 Moreover, No effects of rhizospheric NO on gene expression patterns of putative N transporters and enzymes of glutamine synthesis in European beech, suggesting that these NO effects are mediated by posttranslational modification of proteins.^40,42 More recently, experiments in rice plants reported that NO-mediated increases in the rate of ammonium and nitrate uptake were found to be mediated at the transcriptional level. Surprisingly, NO-induced N uptake in rice cultivars would contribute to nitrogen use efficiency (NUE): i, the induced effect is only observed in high-NUE cultivar rather than in low-NUE cultivar due to that higher NO was only observed in high-NUE plants when supplied by PNN comparison with sole ammonium nutrition; ii, the induced effect can be recorded both in two cultivars when NO level elevated by exogenous application of NO donor.¹⁰ A schematic feedback model is proposed to explain how NO contributes to NUE in rice cultivars: NR-generated NO plays a pivotal role in improving N uptake capacity by increasing root growth and inorganic N uptake, representing a potential strategy for rice adaption to a fluctuating nitrate supply.

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.

Funding

Research in China was funded by the National Natural Science Foundation (No.31672225, 31471936 and 31601821), Innovative Research Team Development Plan of the Ministry of Education of China (No. IRT1256), the 111 Project (No. 12009), and PAPD Project.