Nitrogen (N) availability is a major factor determining plant growth and productivity. Plants acquire inorganic N from the soil, mainly in the form of nitrate and ammonium. To date, researchers have focused on these N sources, and demonstrated that plants exhibit elaborate responses at both physiological and morphological levels. Mixtures of nitrate and ammonium are beneficial in terms of plant growth, as compared to nitrate or ammonium alone, and therefore synergistic responses to both N sources are predicted at different steps ranging from acquisition to assimilation. In this review, we summarize interactions between nitrate and ammonium with respect to uptake, allocation, assimilation, and signaling. Given that cultivated land often contains both nitrate and ammonium, a better understanding of the synergism between these N sources should help to identify targets with the potential to improve crop productivity.

When ammonium is the sole nitrogen (N) source, plant growth is suppressed compared with the situation where nitrate is the N source. This is commonly referred to as ammonium toxicity. It is widely known that a combination of nitrate and ammonium as N source alleviates this ammonium toxicity (nitrate-dependent alleviation of ammonium toxicity), but the underlying mechanisms are still not completely understood. In plants, ammonium toxicity is often accompanied by a depletion of organic acids and inorganic cations, and by an accumulation of ammonium. All these factors have been considered as possible causes for ammonium toxicity. Thus, we hypothesized that nitrate could alleviate ammonium toxicity by lessening these symptoms. We analyzed growth, inorganic N and cation content and various primary metabolites in shoots of Arabidopsis thaliana seedlings grown on media containing various concentrations of nitrate and/or ammonium. Nitrate-dependent alleviation of ammonium toxicity was not accompanied by less depletion of organic acids and inorganic cations, and showed no reduction in ammonium accumulation. On the other hand, shoot growth was significantly correlated with the nitrate concentration in the shoots. This suggests that nitrate-dependent alleviation of ammonium toxicity is related to physiological processes that are closely linked to nitrate signaling, uptake and reduction. Based on transcript analyses of various genes related to nitrate signaling, uptake and reduction, possible underlying mechanisms for the nitrate-dependent alleviation are discussed.

Alternative oxidase (AOX) catalyses the ATP-uncoupling cyanide (CN)-resistant pathway. In this study, our aim was to clarify the physiological role of AOX at low temperature. We examined the effect of low-temperature treatment on CN-resistant respiration (CN-resistant R) and on the transcription of respiratory components in wild-type (WT) and aox1a knock-out transgenic (aox1a) Arabidopsis thaliana plants. In WT leaves, the expression of AOX1a mRNA was strongly induced by the low-temperature treatment, and thus CN-resistant R increased during low-temperature treatment. In aox1a, the CN-sensitive respiration, and the expression of NDB2 and UCP1 were increased compared with WT. We compared several physiological parameters between WT and aox1a. Low-temperature treatment did not result in a visible phenotype to distinguish aox1a from WT. In aox1a, several antioxidant defence genes were induced, and the malondialdehyde content was lower than in WT. Starch content and a ratio of carbon to nitrogen were higher in aox1a than in WT. Our results indicate that a lack of AOX was linked to a difference in the carbon and nitrogen balance, and an up-regulation of the transcription of antioxidant defence system at low temperature. It is likely that AOX is a necessary component in antioxidant defence mechanisms and for the control of a balanced metabolism.

Plants use nitrate, ammonium, and organic nitrogen in the soil as nitrogen sources. Since the elevated CO2 environment predicted for the near future will reduce nitrate utilization by C3 species, ammonium is attracting great interest. However, abundant ammonium nutrition impairs growth, i.e., ammonium toxicity, the primary cause of which remains to be determined. Here, we show that ammonium assimilation by GLUTAMINE SYNTHETASE 2 (GLN2) localized in the plastid rather than ammonium accumulation is a primary cause for toxicity, which challenges the textbook knowledge. With exposure to toxic levels of ammonium, the shoot GLN2 reaction produced an abundance of protons within cells, thereby elevating shoot acidity and stimulating expression of acidic stress-responsive genes. Application of an alkaline ammonia solution to the ammonium medium efficiently alleviated the ammonium toxicity with a concomitant reduction in shoot acidity. Consequently, we conclude that a primary cause of ammonium toxicity is acidic stress.

In order to ensure the cooperative function with the photosynthetic system, the mitochondrial respiratory chain needs to flexibly acclimate to a fluctuating light environment. The non-phosphorylating alternative oxidase (AOX) is a notable respiratory component that may support a cellular redox homeostasis under high-light (HL) conditions. Here we report the distinct acclimatory manner of the respiratory chain to long-and short-term HL conditions and the crucial function of AOX in Arabidopsis thaliana leaves. Plants grown under HL conditions (HL plants) possessed a larger ubiquinone (UQ) pool and a higher amount of cytochrome c oxidase than plants grown under low light conditions (LL plants). These responses in HL plants may be functional for efficient ATP production and sustain the fast plant growth. When LL plants were exposed to short-term HL stress (sHL), the UQ reduction level was transiently elevated. In the wild-type plant, the UQ pool was reoxidized concomitantly with an up-regulation of AOX. On the other hand, the UQ reduction level of the AOXdeficient aox1a mutant remained high. Furthermore, the plastoquinone pool was also more reduced in the aox1a mutant under such conditions. These results suggest that AOX plays an important role in rapid acclimation of the respiratory chain to sHL, which may support efficient photosynthetic performance.

It is widely believed that turnover of nitrogenous (N) compounds (especially proteins) incurs a high respiratory cost. Thus, if protein turnover costs change with temperature, this would influence the dependence of respiration rate on growth temperature. Here, we examined the extent to which protein turnover cost explained differences in N-utilization costs (nitrate uptake/reduction, ammonium assimilation, amino acid and protein syntheses, protein turnover and amino acid export) and in respiration rate with changes in growth temperature. By measurements and literature data, we evaluated each N-utilization cost in Petunia ¥ hybrida petals grown at 20, 25 or 35°C throughout their whole lifespans. Protein turnover cost accounted for 73% of the integrated N-utilization cost on a whole-petal basis at 35°C. The difference in this cost on a dry weight basis between 25 and 35°C accounted for 75% of the difference in N-utilization cost and 45% of the difference in respiratory cost. The cost of nitrate uptake/reduction was high at low growth temperatures. We concluded that respiratory cost in petals was strongly influenced by protein turnover and nitrate uptake/reduction, and on the shoot basis, C investment in biomass was highest at 25°C.

Oxygen uptake rates are increased when concentrated ammonium instead of nitrate is used as sole N source. Several explanations for this increased respiration have been suggested, but the underlying mechanisms are still unclear. To investigate possible factors responsible for this respiratory increase, we measured the O2 uptake rate, activity and transcript level of respiratory components, and concentration of adenylates using Arabidopsis thaliana shoots grown in media containing various N sources. The O2 uptake rate was correlated with concentrations of ammonium and ATP in shoots, but not related to the ammonium assimilation. The capacity of the ATP-coupling cytochrome pathway (CP) and its related genes were up-regulated when concentrated ammonium was sole N source, whereas the ATP-uncoupling alternative oxidase did not influence the extent of the respiratory increase. Our results suggest that the ammonium-dependent increase of the O2 uptake rate can be explained by the up-regulation of the CP, which may be related to the ATP consumption by the plasmamembrane H + -ATPase.

NRT1.1 is a putative nitrate sensor and is involved in many nitrate-dependent responses. On the other hand, a nitrate-independent function of NRT1.1 has been implied, but the clear-cut evidence is unknown. We found that NRT1.1 mutants showed enhanced tolerance to concentrated ammonium as sole N source in Arabidopsis thaliana. This unique phenotype was not observed in mutants of NLP7, which has been suggested to play a role in the nitrate-dependent signaling pathway. Our real-time PCR analysis, and evidence from a literature survey revealed that several genes relevant to the aliphatic glucosinolate-biosynthetic pathway were regulated via a nitrate-independent signal from NRT1.1. When taken together, the present study strongly suggests the existence of a nitrate-independent function of NRT1.1.