The indica and japonica rice (Oryza sativa) subspecies differ in nitrate (NO
) assimilation capacity and nitrogen (N) use efficiency (NUE). Here, we show that a major component of this difference is ...conferred by allelic variation at OsNR2, a gene encoding a NADH/NADPH-dependent NO
reductase (NR). Selection-driven allelic divergence has resulted in variant indica and japonica OsNR2 alleles encoding structurally distinct OsNR2 proteins, with indica OsNR2 exhibiting greater NR activity. Indica OsNR2 also promotes NO
uptake via feed-forward interaction with OsNRT1.1B, a gene encoding a NO
uptake transporter. These properties enable indica OsNR2 to confer increased effective tiller number, grain yield and NUE on japonica rice, effects enhanced by interaction with an additionally introgressed indica OsNRT1.1B allele. In consequence, indica OsNR2 provides an important breeding resource for the sustainable increases in japonica rice yields necessary for future global food security.
The periplasmic (NAP) and membrane-associated (Nar) nitrate reductases of Paracoccus denitrificans are responsible for nitrate reduction under aerobic and anaerobic conditions, respectively. ...Expression of NAP is elevated in cells grown on a relatively reduced carbon and energy source (such as butyrate); it is believed that NAP contributes to redox homeostasis by coupling nitrate reduction to the disposal of excess reducing equivalents. Here, we show that deletion of either
(one of two
homologs in the P. denitrificans genome) or
/
(encoding a bifunctional ppGpp synthetase and hydrolase) eliminates the butyrate-dependent increase in
promoter and NAP enzyme activity. We conclude that ppGpp likely signals growth on a reduced substrate and, together with DksA1, mediates increased expression of the genes encoding NAP. Support for this model comes from the observation that
promoter activity is increased in cultures exposed to a protein synthesis inhibitor that is known to trigger ppGpp synthesis in other organisms. We also show that, under anaerobic growth conditions, the redox-sensing RegAB two-component pair acts as a negative regulator of NAP expression and as a positive regulator of expression of the membrane-associated nitrate reductase Nar. The
and
/
genes are conditionally synthetically lethal; the double mutant has a null phenotype for growth on butyrate and other reduced substrates while growing normally on succinate and citrate. We also show that the second
homolog (
) and
/
have roles in regulation of expression of the flavohemoglobin Hmp and in biofilm formation.
Paracoccus denitrificans is a metabolically versatile Gram-negative bacterium that is used as a model for studies of respiratory metabolism. The organism can utilize nitrate as an electron acceptor for anaerobic respiration, reducing it to dinitrogen via nitrite, nitric oxide, and nitrous oxide. This pathway (known as denitrification) is important as a route for loss of fixed nitrogen from soil and as a source of the greenhouse gas nitrous oxide. Thus, it is important to understand those environmental and genetic factors that govern flux through the denitrification pathway. Here, we identify four proteins and a small molecule (ppGpp) which function as previously unknown regulators of expression of enzymes that reduce nitrate and oxidize nitric oxide.
Nitrate and periplasmic nitrate reductases Sparacino-Watkins, Courtney; Stolz, John F; Basu, Partha
Chemical Society reviews,
01/2014, Letnik:
43, Številka:
2
Journal Article
Recenzirano
Odprti dostop
The nitrate anion is a simple, abundant and relatively stable species, yet plays a significant role in global cycling of nitrogen, global climate change, and human health. Although it has been known ...for quite some time that nitrate is an important species environmentally, recent studies have identified potential medical applications. In this respect the nitrate anion remains an enigmatic species that promises to offer exciting science in years to come. Many bacteria readily reduce nitrate to nitrite
via
nitrate reductases. Classified into three distinct types - periplasmic nitrate reductase (Nap), respiratory nitrate reductase (Nar) and assimilatory nitrate reductase (Nas), they are defined by their cellular location, operon organization and active site structure. Of these, Nap proteins are the focus of this review. Despite similarities in the catalytic and spectroscopic properties Nap from different Proteobacteria are phylogenetically distinct. This review has two major sections: in the first section, nitrate in the nitrogen cycle and human health, taxonomy of nitrate reductases, assimilatory and dissimilatory nitrate reduction, cellular locations of nitrate reductases, structural and redox chemistry are discussed. The second section focuses on the features of periplasmic nitrate reductase where the catalytic subunit of the Nap and its kinetic properties, auxiliary Nap proteins, operon structure and phylogenetic relationships are discussed.
Nitrate in global nitrogen cycle and human health, periplasmic nitrate reductase.
Nitrate reductase (NR) is a key enzyme for nitrogen acquisition by plants, algae, yeasts, and fungi. Nitrate, its main substrate, is required for signaling and is widely distributed in diverse ...tissues in plants. In addition, NR has been proposed as an important enzymatic source of nitric oxide (NO). Recently, NR has been shown to play a role in NO homeostasis by supplying electrons from NAD(P)H through its diaphorase/dehydrogenase domain both to a truncated hemoglobin THB1, which scavenges NO by its dioxygenase activity, and to the molybdoenzyme NO-forming nitrite reductase (NOFNiR) that is responsible for NO synthesis from nitrite. We review how NR may play a central role in plant biology by controlling the amounts of NO, a key signaling molecule in plant cells.
NO synthesis remains a complex picture with many unresolved questions.
NR has been assumed to be the main enzymatic source, but a new and more complex picture for the mechanism of NO synthesis is emerging.
NR, the first enzyme for nitrate assimilation, is a multi-redox protein able to mediate the donation of electrons from NAD(P)H to artificial acceptors and redox proteins. In Chlamydomonas, two of these redox partners are NOFNiR, which efficiently synthesizes NO, and THB1, a truncated hemoglobin, which eliminates NO by its dioxygenase activity.
Homeostasis of the crucial signaling molecule NO in photosynthetic organisms depends on at least two key molybdoenzymes, NR and NOFNiR, as well as on the dioxygenase activity of hemoglobins.
This paper reviews dissimilatory nitrate reduction to ammonium (DNRA) in soils - a newly appreciated pathway of nitrogen (N) cycling in the terrestrial ecosystems. The reduction of NO3− occurs in two ...steps; in the first step, NO3− is reduced to NO2−; and in the second, unlike denitrification, NO2− is reduced to NH4+ without intermediates. There are two sets of NO3−/NO2− reductase enzymes, i.e., Nap/Nrf and Nar/Nir; the former occurs on the periplasmic-membrane and energy conservation is respiratory via electron-transport-chain, whereas the latter is cytoplasmic and energy conservation is both respiratory and fermentative (Nir, substrate-phosphorylation). Since, Nir catalyzes both assimilatory- and dissimilatory-nitrate reduction, the nrfA gene, which transcribes the NrfA protein, is treated as a molecular-marker of DNRA; and a high nrfA/nosZ (N2O-reductase) ratio favours DNRA. Recently, several crystal structures of NrfA have been presumed to producee N2O as a byproduct of DNRA via the NO (nitric-oxide) pathway. Meta-analyses of about 200 publications have revealed that DNRA is regulated by oxidation state of soils and sediments, carbon (C)/N and NO2−/NO3− ratio, and concentrations of ferrous iron (Fe2+) and sulfide (S2−). Under low-redox conditions, a high C/NO3− ratio selects for DNRA while a low ratio selects for denitrification. When the proportion of both C and NO3− are equal, the NO2−/NO3− ratio modulates partitioning of NO3−, and a high NO2−/NO3− ratio favours DNRA. A high S2−/NO3− ratio also promotes DNRA in coastal-ecosystems and saline sediments. Soil pH, temperature, and fine soil particles are other factors known to influence DNRA. Since, DNRA reduces NO3− to NH4+, it is essential for protecting NO3− from leaching and gaseous (N2O) losses and enriches soils with readily available NH4+-N to primary producers and heterotrophic microorganisms. Therefore, DNRA may be treated as a tool to reduce ground-water NO3− pollution, enhance soil health and improve environmental quality.
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•DNRA is a novel and shortest biological pathway of N-cycling in terrestrial ecosystems where NO3- is reduced to NH4+ in soils•This pathway has potential to reduce N2O emissions and protect NO3- from leaching losses•It outcompetes the denitrification process under limited NO3- availability in the system•It enriches soil with NH4+-N and make available to primary producers as N-fertilizer•Meta-analyse revealed that DNRA is mainly regulated by C/N ratio, NO2- / NO3- ratio and S2-
The discovery that dietary (inorganic) nitrate has important vascular effects came from the relatively recent realization of the ‘nitrate‐nitrite‐nitric oxide (NO) pathway’. Dietary nitrate has been ...demonstrated to have a range of beneficial vascular effects, including reducing blood pressure, inhibiting platelet aggregation, preserving or improving endothelial dysfunction, enhancing exercise performance in healthy individuals and patients with peripheral arterial disease. Pre‐clinical studies with nitrate or nitrite also show the potential to protect against ischaemia‐reperfusion injury and reduce arterial stiffness, inflammation and intimal thickness. However, there is a need for good evidence for hard endpoints beyond epidemiological studies. Whilst these suggest reduction in cardiovascular risk with diets high in nitrate‐rich vegetables (such as a Mediterranean diet), others have suggested possible small positive and negative associations with dietary nitrate and cancer, but these remain unproven. Interactions with other nutrients, such as vitamin C, polyphenols and fatty acids may enhance or inhibit these effects. In order to provide simple guidance on nitrate intake from different vegetables, we have developed the Nitrate ‘Veg‐Table’ with ‘Nitrate Units’ each unit being 1 mmol of nitrate (62 mg) to achieve a nitrate intake that is likely to be sufficient to derive benefit, but also to minimize the risk of potential side effects from excessive ingestion, given the current available evidence. The lack of data concerning the long term effects of dietary nitrate is a limitation, and this will need to be addressed in future trials.
Nitrate reductase (NR) is one of the key enzymes for plant nitrogen assimilation and root architecture remodeling. However, crosstalk between NR-mediated signaling and auxin-mediated root development ...in nitrogen-status responses has not been investigated in details before. In this study, root phenotype and auxin distribution in nia1/nia2 (nitrate reductase) double mutant and chl1-5 (nitrate transporter NRT1.1) mutant under different nitrogen availabilities were compared. The nia1/nia2 mutant showed very low expression levels of auxin biosynthetic/signaling genes and was insensitive to nitrogen changes. While the chl1-5 mutant showed a high NR activity with a high level of auxin in the meristematic zone and a weaker response to nitrogen changes, when compared with the wild-type plants. We firstly found that NR activity was roughly positive-correlated with the root auxin level, and there is a crosstalk between nitrate signaling and auxin signaling. The putative signaling pathways downstream of NR have been discussed.
•Nitrate reductase (NR) is a key regulator controlling root system architecture.•NR-deficient mutant showed very low expression levels of auxin biosynthetic/signaling genes.•Nitrate transporter NRT1.1 mutant showed a high NR activity and accumulated a high level of auxin.•NR activity is roughly positive-correlated with the root auxin level.
Plants have evolved adaptive strategies that involve transcriptional networks to cope with and survive environmental challenges. Key transcriptional regulators that mediate responses to environmental ...fluctuations in nitrate have been identified; however, little is known about how these regulators interact to orchestrate nitrogen (N) responses and cell-cycle regulation. Here we report that teosinte branched1/cycloidea/proliferating cell factor1-20 (TCP20) and NIN-like protein (NLP) transcription factors NLP6 and NLP7, which act as activators of nitrate assimilatory genes, bind to adjacent sites in the upstream promoter region of the nitrate reductase gene, NIA1, and physically interact under continuous nitrate and N-starvation conditions. Regions of these proteins necessary for these interactions were found to include the type I/II Phox and Bem1p (PB1) domains of NLP6&7, a protein-interaction module conserved in animals for nutrient signaling, and the histidine- and glutamine-rich domain of TCP20, which is conserved across plant species. Under N starvation, TCP20-NLP6&7 heterodimers accumulate in the nucleus, and this coincides with TCP20 and NLP6&7-dependent up-regulation of nitrate assimilation and signaling genes and down-regulation of the G₂/M cell-cycle marker gene, CYCB1;1. TCP20 and NLP6&7 also support root meristem growth under N starvation. These findings provide insights into how plants coordinate responses to nitrate availability, linking nitrate assimilation and signaling with cell-cycle progression.
Abstract
Nitrate is an important nutrient and signaling molecule in plants, which modulates the expression of many genes and regulates plant growth. In paddy-grown rice (Oryza sativa), nitrogen is ...mostly supplied in the form of ammonium but can also be supplied in the form of nitrate. Several nitrogen transporters and nitrate assimilation enzymes have been identified and functionally characterized in rice. However, little is known regarding the nitrate sensing system in rice, and the regulatory mechanisms of nitrate-related genes remain to be elucidated. In recent years, NIN-like proteins (NLPs) have been described as key transcription factors of nitrogen responses in Arabidopsis thaliana, which implies that OsNLP4 is involved in the regulation of nitrate assimilation and nitrogen use efficiency in rice. Here, we show that OsNLP4 can influence plant growth by affecting nitrate reductase (NR) activity. The growth of OsNLP4 knockdown mutants was reduced when nitrate was supplied, but not when ammonium was supplied. The nitrate concentration was significantly reduced in osnlp4 mutants. Furthermore, the concentrations of iron and molybdenum, essential elements for NR activity, were reduced in OsNLP4 knockdown mutants. We propose that, in addition to the regulation of gene expression within the nitrate signaling pathway, OsNLP4 can affect the NR activity and nitrate-dependent growth of rice. Our results support a working model for the role of OsNLP4 in the nitrate signaling pathway.
The roles of nitrate and nitrite ions as nitric oxide (NO) sources in mammals, complementing NOS enzymes, have recently been the focus of much research. We previously reported that rat skeletal ...muscle serves as a nitrate reservoir, with the amount of stored nitrate being highly dependent on dietary nitrate availability, as well as its synthesis by NOS1 enzymes and its subsequent utilization. We showed that at conditions of increased NO need, this nitrate reservoir is used in situ to generate nitrite and NO, at least in part via the nitrate reductase activity of xanthine oxidoreductase (XOR). We now further investigate the dynamics of nitrate/nitrite fluxes in rat skeletal muscle after first increasing nitrate levels in drinking water and then returning to the original intake level. Nitrate/nitrite levels were analyzed in liver, blood and several skeletal muscle samples, and expression of proteins involved in nitrate metabolism and transport were also measured. Increased nitrate supply elevated nitrate and nitrite levels in all measured tissues. Surprisingly, after high nitrate diet termination, levels of both ions in liver and all muscle samples first declined to lower levels than the original baseline. During the course of the overall experiment there was a gradual increase of XOR expression in muscle tissue, which likely led to enhanced nitrate to nitrite reduction. We also noted differences in basal levels of nitrate in the different types of muscles. These findings suggest complex control of muscle nitrate levels, perhaps with multiple processes to preserve its intracellular levels.
•Nitrate/nitrite levels time course in rat after high nitrate withdrawal was studied.•Nitrate levels returned to baseline in 2-phase nonlinear manner.•Variable nitrate levels in different muscle groups were found.•Nitrate withdrawal was accompanied by changes in several protein expression.
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