Nitrogen assimilation in plants:current status and future prospects

Nitrogen(N)is the driving force for crop yields;however,excessive N application in agriculture not only increases production cost,but also causes severe environmental problems.Therefore,comprehensively understanding the molecular mechanisms of N use efficiency(NUE)and breeding crops with higher NUE is essential to tackle these problems.NUE of crops is determined by N uptake,transport,assimilation,and remobilization.In the process of N assimilation,nitrate reductase(NR),nitrite reductase(Ni R),glutamine synthetase(GS),and glutamine-2-oxoglutarate aminotransferase(GOGAT,also known as glutamate synthase)are the major enzymes.NR and Ni R mediate the initiation of inorganic N utilization,and GS/GOGAT cycle converts inorganic N to organic N,playing a vital role in N assimilation and the final NUE of crops.Besides,asparagine synthetase(ASN),glutamate dehydrogenase(GDH),and carbamoyl phosphate synthetase(CPSase)are also involved.In this review,we summarize the function and regulation of these enzymes reported in three major crops—rice,maize,and wheat,also in the model plant Arabidopsis,and we highlight their application in improving NUE of crops via manipulating N assimilation.Anticipated challenges and prospects toward fully understanding the function of N assimilation and further exploring the potential for NUE improvement are discussed.

Ectopic expression of fungal EcGDH improves nitrogen assimilation and grain yield in rice

NADP（H）-dependent glutamate dehydrogenases（GDH） in lower organisms have stronger ammonium affinity than those in higher plants. Here we report that transgenic rice overexpressing the EcGDH from Eurotium cheralieri exhibited significantly enhanced aminating activities. Hydroponic and field tests showed that nitrogen assimilation efficiency and grain yields were markedly increased in these transgenic plants, especially at the low nitrogen conditions.These results suggest that EcGDH may have potential to be used to improve nitrogen assimilation and grain yield in rice.

Effects of Different Nitrogen and Phosphorus Synergistic Fertilizer on Enzymes and Genes Related to Nitrogen Metabolism in Wheat

In recent years,in order to improve nutrient use efficiency,especially nitrogen use efficiency,fertilizer valueadded technology has been developed rapidly.However,the mechanism of the effect of synergistic fertilizer on plant nitrogen utilization is not clear.A study was,therefore,conducted to explore the activities and gene expression of key enzymes for nitrogen assimilation and the gene expression of nitrogen transporters in wheat after the application of synergistic fertilizer.Soil column experiment was set up in Qingdao Agricultural University experimental base from October 2018 to June 2019.Maleic acid and itaconic acid were copolymerized with acrylic acid as cross-linking monomer to make a fluid gel,which was sprayed on the fertilizer surface to make nitrogen and phosphorus synergistic fertilizer.A total of 6 treatments was set according to different nitrogen and phosphorus fertilizer ratios:(1)100%common nitrogen fertilizer+100%common phosphate fertilizer(2)70%nitrogen synergistic fertilizer+100%phosphorus synergistic fertilizer;(3)100%nitrogen synergistic fertilizer+70%phosphorus synergistic fertilizer;(4)100%nitrogen synergistic fertilizer+100%phosphorus synergistic fertilizer;(5)70%nitrogen synergistic fertilizer+70%phosphorus synergistic fertilizer;(6)100%commercial nitrogen synergistic fertilizer+100%commercial phosphorus synergistic fertilizer.The results are as follows:(1)the enzyme activities of wheat plants under synergistic fertilizer condition were higher than those under ordinary fertilizer,except under the treatment that nitrogen and phosphorus synergistic fertilizer were both reduced;(2)the expression level of the genes under the treatment“100%nitrogen synergistic fertilizer+100%phosphorus synergistic fertilizer”was significantly higher than those in other treatments.Combined with the higher performance of nitrogen concentration in various parts of the plant under the condition of applying synergistic fertilizer,this study indicated that the application of synergistic fertilizer can improve the nitrogen metabolism of the plant by increasing the nitrogen level in the rhizosphere soil,inducing the expression of nitrogen transporter genes and key assimilation enzymes genes.

Overexpression of IbSnRK1 enhances nitrogen uptake and carbon assimilation in transgenic sweetpotato

Nitrogen is an important nutrient for plant development. Nitrogen and carbon metabolisms are tightly linked to physiological functions in plants. In this study, we found that the IbSnRK1 gene was induced by Ca（NO3）2. Its overexpression enhanced nitrogen uptake and carbon assimilation in transgenic sweetpotato. After Ca（^15NO3）2 treatment, the -（15）N atom excess, -（15）N and total N content and nitrogen uptake efficiency（NUE） were significantly increased in the roots, stems, and leaves of transgenic plants compared with wild type（WT） and empty vector control（VC）. After Ca（NO3）2 treatment, the increased nitrate N content, nitrate reductase（NR） activity, free amino acid content, and soluble protein content were found in the roots or leaves of transgenic plants. The photosynthesis and carbon assimilation were enhanced. These results suggest that the IbSnRK1 gene play a important role in nitrogen uptake and carbon assimilation of sweetpotato. This gene has the potential to be used for improving the yield and quality of sweetpotato.

Physiological and Molecular Analysis of Applied Nitrogen in Rice Genotypes

Ten genotypes of rice （Oryza sativa L.） were grown for 30 d in complete nutrient solution with 1 mmol/L （N-insufficient）,4 mmol/L （N-moderate） and 10 mmol/L （N-high） nitrogen levels,and nitrogen efficiency （NE） was analyzed.Growth performance,measured in terms of fresh weight,dry weight and lengths of root and shoot,was higher in N-efficient than in N-inefficient rice genotypes at low N level.Of these 10 genotypes,Suraksha was identified as the most N-efficient,while Vivek Dhan the most N-inefficient.To find out the physiological basis of this difference,the nitrate uptake rate of root and the activities of nitrate assimilatory enzymes in leaves of N-efficient and N-inefficient rice genotypes were studied.Uptake experiments revealed the presence of two separate nitrate transporter systems mediating high-and low-affinity nitrate uptake.Interestingly,the nitrate uptake by the roots of Suraksha is mediated by both high-and low-affinity nitrate transporter systems,while that of Vivek Dhan by only low-affinity nitrate transporter system.Study of the activities and expression levels of nitrate assimilatory enzymes in N-efficient and N-inefficient rice genotypes showed that nitrate reductase （NR） and glutamine synthetase （GS） play important roles in N assimilation under low-nitrogen conditions.

Physiological and Biochemical Mechanisms of Salinity Tolerance in Carex morrowii Boott

Carex species are widely used in many parts of the world and contain a large number of ecologically diverse species.Among the Carex species,some of them are known to be glycophytes,while others are halophytes.Carex morrowii Boott(Cyperaceae)is resistant to trample through their root structure and has an essential ornamental value in the landscape with their leaves.However,no information was found about the level of salinity tolerance/sensitivity of the Carex morrowii among these species.In the present study,changes in trace element contents(Na,K,Ca,Cu,Mn,Mg,Ni,Fe,P,Zn,and N)and their transport from roots to leaves,osmotic regulation,alterations in chlorophyll and carotenoid contents,nitrogen assimilation(nitrate reductase activity;NRA)and total soluble protein content in both roots and leaves of Carex morrowii under different salinity concentrations(50 mM,100 mM,200 mM and 300 mM NaCl)were examined in detail.Our study provides the first detailed data concerning the responses of leaves and roots and the determination of the level of salinity tolerance/sensitivity of the Carex morrowii.The K+/Na+ratio was preserved up to 200 mM NaCl,and accordingly,the element uptake and transport ratios showed that they could control moderate NaCl levels.Ca homeostasis that is maintained even in 200 mM NaCl concentration can be effective in maintaining the structural integrity and selective permeability of the cell membranes,while 300 mM NaCl concentration caused decreased photosynthetic pigments,and deterioration in element content and compartmentation.Moreover,these data suggest that plant parts of Carex morrowii respond differently against varied levels of salinity stress.Although the decrease in NR activity at 200 mM and 300 mM NaCl concentrations in the leaves,NR activity was maintained in the roots.Consequently,Carex morrowii is moderately tolerant to salinity and the carotenoid content and osmotic regulation of Carex morrowii appears to be instrumental in its survival at different salinity levels.Especially the roots of Carex morrowii have a remarkable role in salinity tolerance.

Photosynthesis,Metabolite Composition and Anatomical Structure of Oryza sativa and Two Wild Relatives,O.grandiglumis and O.alta

Photosynthesis, enzyme activities and metabolite pools associated with primary carbon metabolism in leaves were studied in O. grandiglumis and O. alta （wild relatives of rice which produce high biomass） versus O. sativa （a japonica cultivar and a indica-japonica hybrid） to assess their potential for identifying traits which might be utilized to enhance rice productivity. The wild relatives had higher rates of photosynthesis on a fresh weight basis, and higher water use efficiency than the O. sativa lines. There were no striking differences in activities of a number of key enzymes in carbon and nitrogen metabolism between the wild relatives and cultivated rice lines. Along with higher rates of photosynthesis on a fresh weight basis, the leaves of the two wild species had higher nitrate content, higher levels of starch, glucose and fructose, and higher levels of organic acids （malate, succinate and acetate）, compared to the O. sativa lines. The results suggested that O. grandiglumis and O. alta have differences in physiology and primary metabolism which might be exploited to improve growth and productivity of cultivated rice.

Rice Ferredoxin-Dependent Glutamate Synthase Regulates Nitrogen-Carbon Metabolomes and Is Genetically Differentiated between japonica and indica Subspecies

Plants assimilate inorganic nitrogen absorbed from soil into organic forms as Gin and Glu through the glutamine synthetase/glutamine：2-oxoglutarate amidotransferase （GS/GOGAT） cycle. Whereas GS cata- lyzes the formation of Gin from Glu and ammonia, GOGAT catalyzes the transfer of an amide group from Gin to 2-oxoglutarate to produce two molecules of Glu. However, the regulatory role of the GS/GOGAT cycle in the carbon-nitrogen balance is not well understood. Here, we report the functional characterization of rice ABNORMAL CYTOKININ RESPONSE 1 （ABC1） gene that encodes a ferredoxin-dependent （Fd）- GOGAT. The weak mutant allele abcl-1 mutant shows a typical nitrogen-deficient syndrome, whereas the T-DNA insertional mutant abcl-2 is seedling lethal. Metabolomics analysis revealed the accumulation of an excessive amount of amino acids with high N/C ratio （Gin and Asn） and several intermediates in the tricarboxylic acid cycle in abcl-1, suggesting that ABC1 plays a critical role in nitrogen assimilation and carbon-nitrogen balance. Five non-synonymous single-nucleotide polymorphisms were identified in the ABC1 coding region and characterized as three distinct haplotypes, which have been highly and specifically differentiated between japonica and indica subspecies. Collectively, these results suggest that ABC1/ OsFd-GOGAT is essential for plant growth and development by modulating nitrogen assimilation and the carbon-nitrogen balance.

Responses of photosynthetic characteristics and enzyme activity of nitrogen metabolism to low nitrogen in maize with different nitrogen tolerance

Understanding the physiological processes associated with leaf photosynthetic characteristics and nitrogen(N)assimilation during grain-filling stage are helpful for enhancing nitrogen utilization efficiency(NUtE)of maize.In this study,the leaf photosynthetic and N assimilation parameters in maize,including Zhengdan 958(ZD958),a low-N tolerance cultivar and Huanong 138(HN138),a low-N sensitive cultivar under different N rates were examined.Results showed that ZD958 displayed significant increases on grain yield and NUtE than that in HN138.Analyses on the leaf photosynthetic and N assimilation-associated processes indicated that ZD958 had higher leaf N remobilization(Rem N),net photosynthetic rate(Pn)and photosynthetic N use efficiency(PNUE)with respect to those of HN138 during grain-filling stage.In addition,ZD958 was also shown to be higher activities of leaf nitrate reductase(NR),glutamine synthetase(GS),nitrate reductase(GDH)and glutamine synthetase(GAGOT)than those of HN138.The leaf PNUE was significantly positively correlated with NR,GS,GDH,GOGAT suggesting that leaf PNUE and NR,GS,GDH,GOGAT jointly determined the N remobilization efficiency and the leaf N remobilization during post-silking.These results suggested that ZD958 possessed improved PNUE,NR and GS activities in leaves during grain-filling stage that contributes improve grain weights and yield formation capacities upon under low-N conditions.