Ammonium Assimilation Research Articles

Comparative studies on the response of Zostera marina leaves and roots to ammonium stress and effects on nitrogen metabolism.

Coastal eutrophication has resulted in the rapid loss and deterioration of seagrass beds worldwide. The high concentration of ammonium in eutrophic aquatic environments has been invoked as the main cause. In this study, leaves and roots of the seagrass Zostera marina were treated with simulated eutrophic seawater with elevated ammonium concentrations. The tolerance to ammonium stress and mechanism of nitrogen metabolism detoxification in different tissues were investigated. The results showed that high ammonium stress significantly affected the growth of leaves and had a negative effect on photosynthesis. The root activity of Z. marina was not inhibited at ammonium concentrations of ≤100mg/L, indicating that the roots exhibited tolerance to ammonium stress. Increasing ammonium concentrations led to a higher increase of ammonium and free amino acid (FAA) contents in leaves than in roots. However, nitrogen storage decreased in Z. marina leaves after high ammonium treatments. The enzyme activity and gene expression of glutamine synthetase (GS) in roots were significantly higher than in the leaves even under ammonium stress. Meanwhile, ammonium stress increased the enzyme activities and gene expression of glutamate synthase (GOGAT) and glutamate dehydrogenase (GDH) in roots, which suggested that the roots had a strong ability to assimilate ammonium under ammonium stress. In contrast, although the GOGAT and GDH activity and gene expression in the leaves were initially increased, they significantly decreased when the ammonium concentration exceeded 100mg/L. These results indicated that the concentration of 100mg/L might be a threshold marking a transition from tolerance to toxicity for the leaves. Our study demonstrates that Z. marina leaves could be prone to higher damage than roots because the mechanism of ammonium assimilation in leaves is more susceptible to ammonium toxicity.

Analysis of the Traits of Nitrogen Metabolism Pathways for Several Forest Soils in Eastern China

Nitrogen metabolism pathways mediated by microorganisms play an important role in maintaining the structure and functional stability of soil ecosystems. Clarifying the relationships between microbial communities and nitrogen metabolism pathways can expand our understanding of nitrogen metabolism pathways at a microscopic level. However, the horizontal gene transfer of microorganisms means that taxonomy-based methods cannot be easily applied. A growing number of studies have shown that functional traits affect community construction and ecosystem functions. Using methods based on functional traits to study soil microbial communities can, therefore, better characterize nitrogen metabolism pathways. Here, five typical forest soils in China, namely black soil(Harbin, Heilongjiang), dark-brown earth(Changbaishan, Jilin), yellow-brown earth(Wuhan, Hubei), red earth(Fuzhou, Fujian), and humid-thermo ferralitic soil(Ledong, Hainan), were selected to study the traits of nitrogen metabolism pathways using metagenomic technology combined with the trait-based methods. The studied nitrogen metabolism pathways were ammonia assimilation, nitrate dissimilatory reduction, nitrate assimilatory reduction, denitrification, nitrification, nitrogen fixation, and anaerobic ammonia oxidation. The results showed that bacteria dominated the metagenomic library, accounting for 98.02% of all the sequences. Across all domains, the most common pathway was ammonia assimilation. For example, an average of 2830 ammonia assimilation pathway genes were detected for every million annotated bacterial sequences. In comparison, nitrogen fixation and anaerobic ammonia oxidation were the least detected pathways, accounting for 28.3 and 10.7 per million sequences, respectively. Different microorganisms can participate in a same nitrogen metabolism pathway, and the community structure of different soils was variable. The five typical forest soils in China show the same microbial nitrogen metabolism pathway traits; however, the community structure of the microorganisms mediating these processes was found to vary.

Genome-Wide Analyses of Proteome and Acetylome in Zymomonas mobilis Under N2-Fixing Condition.

Zymomonas mobilis, a promising candidate for industrial biofuel production, is capable of nitrogen fixation naturally without hindering ethanol production. However, little is known about the regulation of nitrogen fixation in Z. mobilis. We herein conducted a high throughput analysis of proteome and protein acetylation in Z. mobilis under N2-fixing conditions and established its first acetylome. The upregulated proteins mainly belong to processes of nitrogen fixation, motility, chemotaxis, flagellar assembly, energy production, transportation, and oxidation–reduction. Whereas, downregulated proteins are mainly related to energy-consuming and biosynthetic processes. Our acetylome analyses revealed 197 uniquely acetylated proteins, belonging to major pathways such as nitrogen fixation, central carbon metabolism, ammonia assimilation pathway, protein biosynthesis, and amino acid metabolism. Further, we observed acetylation in glycolytic enzymes of central carbon metabolism, the nitrogenase complex, the master regulator NifA, and the enzyme in GS/GOGAT cycle. These findings suggest that protein acetylation may play an important role in regulating various aspects of N2-metabolism in Z. mobilis. This study provides new knowledge of specific proteins and their associated cellular processes and pathways that may be regulated by protein acetylation in Z. mobilis.

Drought, Salinity, and Low Nitrogen Differentially Affect the Growth and Nitrogen Metabolism of Sophora japonica (L.) in a Semi-Hydroponic Phenotyping Platform.

Abiotic stresses, such as salinity, drought, and nutrient deficiency adversely affect nitrogen (N) uptake and assimilation in plants. However, the regulation of N metabolism and N pathway genes in Sophora japonica under abiotic stresses is unclear. Sophora japonica seedlings were subjected to drought (5% polyethylene glycol 6,000), salinity (75mM NaCl), or low N (0.01mM NH4NO3) for 3weeks in a semi-hydroponic phenotyping platform. Salinity and low N negatively affected plant growth, while drought promoted root growth and inhibited aboveground growth. The NH4+/NO3− ratio increased under all three treatments with the exception of a reduction in leaves under salinity. Drought significantly increased leaf NO2− concentrations. Nitrate reductase (NR) activity was unaltered or increased under stresses with the exception of a reduction in leaves under salinity. Drought enhanced ammonium assimilation with increased glutamate synthase (GOGAT) activity, although glutamine synthetase (GS) activity remained unchanged, whereas salinity and low N inhibited ammonium assimilation with decreased GS activity under salt stress and decreased GOGAT activity under low N treatment. Glutamate dehydrogenase (GDH) activity also changed dramatically under different stresses. Additionally, expression changes of genes involved in N reduction and assimilation were generally consistent with related enzyme activities. In roots, ammonium transporters, especially SjAMT1.1 and SjAMT2.1a, showed higher transcription under all three stresses; however, most nitrate transporters (NRTs) were upregulated under salinity but unchanged under drought. SjNRT2.4, SjNRT2.5, and SjNRT3.1 were highly induced by low N. These results indicate that N uptake and metabolism processes respond differently to drought, salinity, and low N conditions in S. japonica seedlings, possibly playing key roles in plant resistance to environmental stress.

Mechanistic insights into ureolysis mediated calcite precipitation

Microbially induced calcite precipitation (MICP) has applications in self-healing concrete, soil consolidation and bio-grouting. Development of models of processes involved in MICP is a step towards gaining mechanistic insights and building reliable predictive models of MICP. This work deals with understanding the reactions and transport processes involved in ureolysis mediated calcite precipitation in Sporosarcina pasteurii (S. pasteurii). Particularly, a simple structured model that describes urea uptake, ureolysis, ammonium assimilation and ammonium excretion processes is developed. The structured model describes extra-cellular processes like transport processes and important intra-cellular reactions involved in ureolysis. The model parameters are estimated using measured concentrations of extracellular and intracellular metabolites during experiments. In this work, The maximum specific growth rate and yield coefficient of S. pasteurii are found to be 0.78 d−1 and 4.93 gDW/g, respectively. The maximum urease activity of 1.79 μmoles/min/gDW biomass was observed during the fourth day of growth when grown in minimal media. It is shown that the urea uptake can be described by a first-order kinetics suggesting active transport. The estimated parameter of rate constant of first-order kinetics is 9.8 × 10−2d−1. On the other hand, the ammonium ion excretion takes place through free diffusion. The estimated parameter of permeability coefficient for ammonium ion diffusion is 9.2 × 10−3 m d−1.

Impacts of environmental conditions, and allelic variation of cytosolic glutamine synthetase on maize hybrid kernel production

Cytosolic glutamine synthetase (GS1) is the enzyme mainly responsible of ammonium assimilation and reassimilation in maize leaves. The agronomic potential of GS1 in maize kernel production was investigated by examining the impact of an overexpression of the enzyme in the leaf cells. Transgenic hybrids exhibiting a three-fold increase in leaf GS activity were produced and characterized using plants grown in the field. Several independent hybrids overexpressing Gln1-3, a gene encoding cytosolic (GS1), in the leaf and bundle sheath mesophyll cells were grown over five years in different locations. On average, a 3.8% increase in kernel yield was obtained in the transgenic hybrids compared to controls. However, we observed that such an increase was simultaneously dependent upon both the environmental conditions and the transgenic event for a given field trial. Although variable from one environment to another, significant associations were also found between two GS1 genes (Gln1-3 and Gln1-4) polymorphic regions and kernel yield in different locations. We propose that the GS1 enzyme is a potential lead for producing high yielding maize hybrids using either genetic engineering or marker-assisted selection. However, for these hybrids, yield increases will be largely dependent upon the environmental conditions used to grow the plants.

Cr (VI)-induced oxidative damage impairs ammonia assimilation into organic forms in Solanum lycopersicum L.

Chromium (Cr) is a dangerous metal that has been found in the environment in high amounts as the result of numerous human activities, with proven negative effects on various organisms, including plants. The exposure of some plant species to this metal resulted in reduced growth, impaired nutrient uptake, and led to an overproduction of reactive oxygen species (ROS), ultimately causing oxidative damage. Nevertheless, studies regarding the impact of Cr on nitrogen (N) metabolism, the main limiting factor for plant growth, either directly or through the occurrence of oxidative stress, are scarce. Thus, this work aimed to explore the interplay between ammonia (NH4+) assimilation, redox homeostasis and antioxidant (AOX) system of tomato plants exposed to increased concentrations of Cr (VI) (0, 5 and 10 µM). The results revealed that Cr (VI) induced biometric damages, with roots being the most affected organ. Glutamine synthetase (GS) was differentially influenced by the metal at the gene expression, protein and activity levels, whereas glutamate dehydrogenase (GDH) activity was enhanced. Moreover, in Cr (VI)-treated plants, the observed oxidative damage was dependent on metal concentration and plant organ, with a differential accumulation of ROS and a generalized increase of lipid peroxidation (LP) recorded, being paired with an overall failure of the enzymatic antioxidant (AOX) system. Still, the non-enzymatic AOX component had a relevant role in the protection of plants against Cr (VI). Overall, the obtained results suggest that Cr (VI)-mediated plant growth reduction resulted from the occurrence of oxidative stress, coupled with the lack of enzymatic AOX defense, which, consequently, affected the NH4+ assimilatory route.

Redox-Controlled Ammonium Storage and Overturn in Ediacaran Oceans

As a key nutrient, nitrogen can limit primary productivity and carbon cycle dynamics, but also evolutionary progress. Given strong redox-dependency of its molecular speciation, environmental conditions can control nitrogen localization and bioavailability. This particularly applies to periods in Earth history with strong and frequent redox fluctuations, such as the Neoproterozoic. We here report on chlorophyll-derived porphyrins and maleimides in Ediacaran sediments from Oman. Exceptionally light δ15N values (< –10‰) in maleimides derived from anoxygenic phototrophs point towards ammonium assimilation at the chemocline, whereas the isotopic offset between kerogens and chlorophyll-derivatives indicates a variable regime of cyanobacterial and eukaryotic primary production in surface waters. Biomarker and maleimide mass balance considerations imply shallow euxinia during the terminal Ediacaran and a stronger contribution of anoxygenic phototrophs to primary productivity, possibly as a consequence of nutrient ‘lockup’ in a large anoxic ammonium reservoir. Synchronous δ13C and δ15N anomalies at the Ediacaran–Cambrian boundary may reflect one in a series of overturn events, mixing ammonium and isotopically-light DIC into oxic surface waters. By modulating access to nitrogen, environmental redox conditions may have periodically affected Ediacaran primary productivity, carbon cycle perturbations, and possibly played a role in the timing of the metazoan radiation across the terminal Ediacaran and early Cambrian.

MdUGT88F1-mediated phloridzin biosynthesis coordinates carbon and nitrogen accumulation in apple.

The high accumulation of phloridzin makes apple (Malus domestica) unique in the plant kingdom, which suggests a vital role of its biosynthesis in physiological processes. In our previous study, silencing MdUGT88F1 (a key UDP-GLUCOSE: PHLORETIN 2'-O-GLUCOSYLTRANSFERASE gene) revealed the importance of phloridzin biosynthesis in apple development and Valsa canker resistance. Here, results from MdUGT88F1-silenced lines showed that phloridzin biosynthesis was indispensable for normal chloroplast development and photosynthetic carbon fixation by maintaining MdGLK1/2 (GOLDEN2-like1/2) expression. Interestingly, increased phloridzin biosynthesis did not affect plant (or chloroplast) development, but reduced nitrogen accumulation, leading to chlorophyll deficiency, light sensitivity, and sugar accumulation in MdUGT88F1-overexpressing apple lines. Further analysis revealed that MdUGT88F1-mediated phloridzin biosynthesis negatively regulated the cytosolic glutamine synthetase1-asparagine synthetase-asparaginase (GS1-AS-ASPG) pathway of ammonium assimilation and limited chlorophyll synthesis in apple shoots. The interference of phloridzin biosynthesis in the GS1-AS-ASPG pathway was also assumed to be associated with its limitation of the carbon skeleton of ammonium assimilation through metabolic competition with the tricarboxylic acid cycle. Taken together, our findings shed light on the role of MdUGT88F1-mediated phloridzin biosynthesis in the coordination between carbon and nitrogen accumulation in apple trees.

Physiological characteristics and miRNA sequencing of two root zones with contrasting ammonium assimilation patterns in Populus.

The net ammonium fluxes differ among the different root zones of Populus, but the physiological and microRNA regulatory mechanisms are unclear. To elucidate the physiological and miRNA regulatory mechanisms, we investigated the two root zones displaying significant differences in net NH4+ effluxes of P. × canescens. Populus plantlets were cultivated with 500μM NH4Cl for 10days. Six plants were randomly selected to determine the net NH4+ fluxes using a noninvasive microtest technique. High-throughput sequencing were used to determine the dynamic expression profile of miRNA among the different root zones of Populus. Net NH4+ efflux in zone I (from 0 to 40mm from the root apex) was - 19.64pmolcm-2s-1 and in zone II (from 40 to 80mm) it was - 43.96pmolcm-2s-1. The expression of eleven miRNAs was significantly upregulated, whereas fifteen miRNAs were downregulated. Moreover, eighty-eight target genes of the significantly differentially expressed miRNAs were identified in root zone II compared with zone I. Particularly, ptc-miR171a/b/e and their target, SCL6, were found to be important for the difference in net NH4+ effluxes in the two root zones. Moreover, the expression of the target of ptc-miR169d, NFYA3 was upregulated in root zone II compared with root zone I, contributing to increased NH4+ efflux and decreased NH4+ assimilation in root zone II. These results indicate that miRNAs regulate the expression levels of their target genes and thus play key roles in net NH4+ fluxes and NH4+ assimilation in different poplar root zones.

Glutamate dehydrogenase isogenes CsGDHs cooperate with glutamine synthetase isogenes CsGSs to assimilate ammonium in tea plant (Camellia sinensis L.)

Glutamate dehydrogenase (GDH) is a central enzyme in nitrogen metabolism, assimilating ammonia into glutamine or deaminating glutamate into α-oxoglutarate. Tea (Camellia sinensis L.) plants assimilate ammonium efficiently, but the role of CsGDH in ammonium assimilation remains unclear. We confirmed that tea has three GDH isogenes: CsGDH1-3. Bioinformatic analysis showed that CsGDH1 encodes the β-GDH subunit, CsGDH2/3 encode the α-GDH subunit, and their proteins all feature an NADH-specific motif. CsGDH1 is mainly expressed in mature leaves and roots, CsGDH3 is mainly expressed in new shoots and roots, and CsGDH2 has the highest expression level in flowers compared to the other five tissues. Expression patterns of CsGDHs and glutamine synthetase isogenes (CsGSs) under different ammonium concentrations suggested that CsGDHs cooperate with CsGSs to assimilate ammonium, especially under high ammonium conditions. Inhibition of GS and its isogenes resulted in significant induction of CsGDH3 in roots and CsGDH2 in leaves, indicating their potential roles in ammonium assimilation. Moreover, CsGDHs transcripts were highly abundant in chlorotic tea leaves, in constrast to those of CsGSs, suggesting that CsGDHs play a vital role in ammonium assimilation in chlorotic tea mutant. Altogether, our circumstantial evidence that CsGDHs cooperate with CsGSs in ammonium assimilation provides a basis for unveiling their functions in tea plants.

Excessive ammonium assimilation by plastidic glutamine synthetase causes ammonium toxicity in Arabidopsis thaliana

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.

Biotic and Isotopic Vestiges of Oligotrophy on Continental Shelves During Oceanic Anoxic Event 2

AbstractThe widespread expansion of the oxygen minimum zone onto shelves has been commonly regarded as a primary cause of benthos extinction in epicratonic sea ecosystems during the Cenomanian–Turonian boundary event (CTBE). However, neither lithology, geochemical proxies, nor micropaleontological data support this hypothesis. Instead, our integrated foraminiferal and dinoflagellate cyst study, corroborated by δ13Corg and δ15Norg data, indicate that the biota were impacted by an abrupt shift to well oxygenated oligotrophic conditions and a collapse of primary productivity in the epicontinental Central European Basin. Because the event was concurrent with the development of extensive and extreme oceanic bottom water anoxia that reached the photic zone in oceanic settings, we infer that the biotic crisis in the shelf seas during Oceanic Anoxic Event 2 (OAE2), and possibly during other OAEs, was triggered by this anomalous nutrient cycling in Earth's oceans. This phenomenon was presumably associated with intensive denitrification combined with anammox activity in the deep “ammonium oceans,” which caused a significant loss of biologically reactive nitrogen from the ocean system. Impingement of ammonium‐rich anoxic waters on the photic zone resulted in primary productivity based primarily on ammonium assimilation, as recorded by strongly 15N‐depleted organic matter deposited in the oceans during the CTBE. We propose that, unlike in the oceanic settings, productivity in the well‐oxygenated, oligotrophic epicontinental seas was nitrate‐based, as evidenced by strongly 15N‐enriched organic matter deposited in the contemporaneous epicontinental sea. These very high δ15Norg values (>+5‰) were related to the spreading of shallow oceanic waters carrying 15N‐enriched nitrate onto epicontinental settings.

English

Nitrogen (N), one of the most important elements for plant growth, is needed by plants in large quantities. However, this nutrient has limited supply in the soil. Arbuscular mycorrhizal fungi (AMF) are known for their ability to form symbiotic association with plants and transfer the mineral nutrients to the host plants. To validate this hypothesis on sorghum plants, three ecotypes of this cereal (3p4, 3p9 and 4p11) were cultivated with and without AMF under low nitrogen concentration (0.5 mM NH4+). Growth parameters were determined and key enzymes responsible for nitrogen and carbon metabolisms such as glutamine synthetase (GS), glutamate dehydrogenase (GDH), phosphoenolpyruvate carboxylase (PEPC), isocitrate dehydrogenase (ICDH), malate dehydrogenase (MDH) and asparate aminotransferase (AAT) were measured. For the three sorghum ecotypes, mycorrhizal plants showed a higher plant growth compared to the control plants. The biochemical parameters revealed a significant increase in the nitrogen assimilatory enzymes; GS and GDH in the leaves and roots of mycorrhizal plants. Furthermore, mycorrhizal fungi also appear to have a significant effect on carbon assimilatory enzymes. These enzymes are known to have a cardinal role in the provision of carbon skeletons essential for the assimilation of ammonium and thus, amino acids synthesis. Our study indicates clearly that AMF can be an efficient way to optimize nitrogen uptake and/or assimilation by plants and thus improve the crop yields with lower amount of nitrogen fertilizers. © 2021 Friends Science Publishers

Simultaneous removal of phosphorous and nitrogen by ammonium assimilation and aerobic denitrification of novel phosphate-accumulating organism Pseudomonas chloritidismutans K14

Pseudomonas chloritidismutans K14, a novel phosphate-accumulating organism with the capacity to perform ammonium assimilation, aerobic denitrification, and phosphorus removal, was isolated from aquaculture sediments. It produced no hemolysin, and showed susceptibility to most antibiotics. Optimum conditions were achieved with sodium pyruvate as a carbon source, a C/N ratio of 10, pH of 7.5, temperature of 27 °C, P/N ratio of 0.26, and shaking at 140 rpm. Under optimum conditions, the highest removal efficiencies of ammonium, nitrite, and nitrate were 99.82%, 99.11%, and 99.78%, respectively; the corresponding removal rates were 6.27, 4.51, and 4.99 mg/L/h. The strain removed over 98% of phosphorus, and over 87% of chemical oxygen demand. The highest biomass nitrogen during ammonium assimilation was 99.18 mg/L; no gaseous nitrogen was produced. The genes involved in nitrogen and phosphorus removal were amplified by PCR. This study demonstrated the potential application prospects of strain K14 for nitrogen and phosphorus removal.

Rice amino acid transporter‐like 6 (OsATL6) is involved in amino acid homeostasis by modulating the vacuolar storage of glutamine in roots

Glutamine is a product of ammonium (NH4 + ) assimilation catalyzed by glutamine synthetase (GS) and glutamate synthase (GOGAT). The growth of NH4 + -preferring paddy rice (Oryza sativa L.) depends on root NH4 + assimilation and the subsequent root-to-shoot allocation of glutamine; however, little is known about the mechanism of glutamine storage in roots. Here, using transcriptome and reverse genetics analyses, we show that the rice amino acid transporter-like6 (OsATL6) protein exports glutamine to the root vacuoles under NH4 + -replete conditions. OsATL6 was expressed, along with OsGS1;2 and OsNADH-GOGAT1, in wild-type (WT) roots fed with sufficient NH4 Cl, and was induced by glutamine treatment. We generated two independent Tos17 retrotransposon insertion mutants showing reduced OsATL6 expression to determine the function of OsATL6. Compared with segregants lacking the Tos17 insertion, the OsATL6 knock-down mutant seedlings exhibited lower root glutamine content but higher glutamine concentration in the xylem sap and greater shoot growth under NH4 + -replete conditions. The transient expression of monomeric red fluorescent protein-fused OsATL6 in onion epidermal cells confirmed the tonoplast localization of OsATL6. When OsATL6 was expressed in Xenopus laevis oocytes, glutamine efflux from the cell into the acidic bath solution increased. Under sufficient NH4 + supply, OsATL6 transiently accumulated in sclerenchyma and pericycle cells, which are located adjacent to the Casparian strip, thus obstructing the apoplastic solute path, and in vascular parenchyma cells of WT roots before the peak accumulation of GS1;2 and NADH-GOGAT1 occurred. These findings suggest that OsATL6 temporarily stores excess glutamine, produced by NH4 + assimilation, in root vacuoles before it can be translocated to the shoot.

Integrative Transcriptomic and Proteomic Analysis Reveals an Alternative Molecular Network of Glutamine Synthetase 2 Corresponding to Nitrogen Deficiency in Rice (Oryza sativa L.).

Nitrogen (N) is an essential nutrient for plant growth and development. The root system architecture is a highly regulated morphological system, which is sensitive to the availability of nutrients, such as N. Phenotypic characterization of roots from LY9348 (a rice variety with high nitrogen use efficiency (NUE)) treated with 0.725 mM NH4NO3 (1/4N) was remarkable, especially primary root (PR) elongation, which was the highest. A comprehensive analysis was performed for transcriptome and proteome profiling of LY9348 roots between 1/4N and 2.9 mM NH4NO3 (1N) treatments. The results indicated 3908 differential expression genes (DEGs; 2569 upregulated and 1339 downregulated) and 411 differential abundance proteins (DAPs; 192 upregulated and 219 downregulated). Among all DAPs in the proteome, glutamine synthetase (GS2), a chloroplastic ammonium assimilation protein, was the most upregulated protein identified. The unexpected concentration of GS2 from the shoot to the root in the 1/4N treatment indicated that the presence of an alternative pathway of N assimilation regulated by GS2 in LY9348 corresponded to the low N signal, which was supported by GS enzyme activity and glutamine/glutamate (Gln/Glu) contents analysis. In addition, N transporters (NRT2.1, NRT2.2, NRT2.3, NRT2.4, NAR2.1, AMT1.3, AMT1.2, and putative AMT3.3) and N assimilators (NR2, GS1;1, GS1;2, GS1;3, NADH-GOGAT2, and AS2) were significantly induced during the long-term N-deficiency response at the transcription level (14 days). Moreover, the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis demonstrated that phenylpropanoid biosynthesis and glutathione metabolism were significantly modulated by N deficiency. Notably, many transcription factors and plant hormones were found to participate in root morphological adaptation. In conclusion, our study provides valuable information to further understand the response of rice roots to N-deficiency stress.

Ammonia reduction by the gdhA and glnA genes from bacteria in laying hens

Ammonia emissions are a high-focus pollution issue in the livestock industry. Ammonia-degrading bacteria can assimilate ammonia nitrogen as a nitrogen source to promote their growth and reproduction, providing an environmentally friendly, low-cost and safe biological way to reduce ammonia emissions from livestock. However, it remains unclear how ammonia-degrading bacteria reduce ammonia emissions from animals and what are the key ammonia assimilation genes. In the present study, two strains with ammonia nitrogen-degrading abilities (Enterococcus faecium strain C2 and Bacillus coagulans strain B1) were screened from laying chicken caecal and faecal samples and reduced ammonia emission rates by 53.60% and 31.38%, respectively. The expression levels of the ammonia assimilation genes gdhA, glnA, and GMPS increased significantly. On this basis, we successfully constructed three clone strains (PET-GDH, PET-GS, and PET-GMPS) that expressed the gdhA, glnA and GMPS genes in E. coli, respectively, to verify their ammonia-reducing activities. The results of an in vitro fermentation study showed that the ammonia production of the PET-GDH and PET-GS groups was significantly lower than that of the empty vector group (p < 0.05), with ammonia emission reduction rates of 55.5% and 54.8%, respectively. However, there was no difference between the PET-GMPS and empty vector groups. These results indicate that gdhA and glnA may be key genes involved in the bacterial-mediated regulation of ammonia emissions by laying hens, and ammonia emissions may be reduced by regulating their expression. The results of the present study provide a theoretical basis for the construction of engineered bacteria to reduce ammonia production in animals.

Divergence of plastid 2-oxoglutarate “only” transporters away from general transporters by using a cysteine-rich architecture

The common carbon and nitrogen currency, 2-oxoglutarate, could become a valuable resource for nitrogen assimilation and carbon centered biochemical fates. Here in this in silico study, a myriad of factors was used, namely phylogeny, sequence comparisons, and presence and location of clustered cysteines in specific plastid transporters of 2-oxoglutarate, to examine their evolution away from more generalized transporters. This transition would be to adopt the capability of internalizing 2-oxoglutarate alone or with superior specificities at the expense of malate. In phylogeny, the specific 2-oxoglutarate transporters (Cluster 1) are clustered in a separate clade away from 2 clades of general transporters (Cluster 2 and 3). The exclusivity (Cluster 1) and promiscuity of transporters (Cluster 2 and 3) compared to Arabidopsis counterparts characterized prior to this study, were used as a benchmark for my study. Within this mother clade of exclusive transporters, C4 and C3 2-oxoglutarate transporters once again form separate clusters of monophyly. Furthermore, a pattern of Cys –X-X-Cys-X(19)-Cys is conserved within the 2-oxo-glutarate only transporters that is missing in general transporters. Cysteines which are functionally key residues are inferred to be mediating intra- or inter-reactive disulfide bond formation or using a thiol (sulfhydryl) group for transport or to be forming a metal binding site. When a disulfide bond prediction tool was employed, it showed with negligible doubt that the Cys-X-X Cys-X(19) -Cys region was a strong contender for 2 separate disulfide bonds, although the middle cysteine was predicted to be involved in both. In addition, Cluster 2 general Zea mays C4 transporters are shown to be more recalcitrant to mutations of cysteines, compared to Panicum and Oryza counterparts. The study of 2-oxoglutarate and its availability in the chloroplast could play a two-prong role in C4 plants: to be a candidate for synthesis of bundle sheath cell Rubisco enzyme, which makes up ~50% of plant proteins, via ammonia assimilation, and even playing a role in carbon-centered biochemical pathways. This study could greatly facilitate choices in the tinkering of the right transporters for a future C4 rice in a climate change impacted world.

Constraining nitrification by intermittent aeration to achieve methane-driven ammonia recovery of the mainstream anaerobic effluent

Mainstream anaerobic treatment has the potential to capture organic energy, and represents a sustainable development trend, but with the problems of low biogas quality and dissolved methane emissions. In this study, methane-driven ammonia recovery of anaerobic effluent was proposed. A 380-day long-term experiment, which was divided into four phases according to different aeration modes, was conducted. The ammonia conversion and microbial characteristics shows that ammonia oxidizing bacteria (AOB) were constrained during Phases 2 (DO: <0.2 mg L−1) and 4 (DO: 0.1–1.6 mg L−1), and were active during Phase 3 (DO: 2–4 mg L−1). During phase 4, when the intermittent aeration was used, the total nitrogen removal rate was higher than during Phases 2 and 3, and nearly 100% ammonia was removed. Methylomonas, a genus of methane oxidizing bacteria (MOB), was enriched during Phase 4. The serum bottle experiment confirmed that the ammonia removal occurred through the MOB assimilation. The protein content in the CH4-added group was 35.5%, which was higher than in the group without CH4 (23.3%). The powerful ammonia assimilation and protein synthesis capabilities of MOB give a meaning to the anaerobic effluent for ammonia recovery and protein production. Intermittent aeration could be used to constrain AOB and improve ammonia recovery efficiency.