Nitrogen metabolism of Plantago lanceolata as dependent on the supply of mineral nutrients

Various aspects of nitrogen metabolism of Plantago lanceolata L., a grassland species from a relatively nutrient poor habitat, were investigated under high nutrient conditions or low nutrient conditions. In addition the responses after switching the plants from high nutrient conditions to low nutrient conditions and vice versa were studied. The activities of nitrate reductase (NR), glutamate dehydrogenase (GDH) and glutamine synthetase (GS) were determined. In the roots they were correlated with the level of the nutrient supply. Not only NR but also GDH and GS increased upon a switch from low nutrients to high nutrients and decreased upon a switch from high nutrients to low nutrients. Reduced nitrogen content of both roots and shoots was also correlated with the nutrient supply and changed rapidly after a switch. The flexibility of the nitrogen metabolism of Plantago lanceolata is discussed in relation to its ecological habitat.     

Over-expression of cytosolic glutamine synthetase increases photosynthesis and growth at low nitrogen concentrations.

Nitrogen, which is a major limiting nutrient for plant growth, is assimilated as ammonium by the concerted action of glutamine synthetase (GS) and glutamate synthase (GOGAT). GS catalyses the critical incorporation of inorganic ammonium into the amino acid glutamine. Two types of GS isozymes, located in the cytosol (GS1) and in the chloroplast (GS2) have been identified in plants. Tobacco (Nicotiana tabacum) transformants, over-expressing GS1 driven by the constitutive CaMV 35S promoter were analysed. GS in leaves of GS-5 and GS-8 plants was up-regulated, at the level of RNA and proteins. These transgenic plants had six times higher leaf GS activity than controls. Under optimum nitrogen fertilization conditions there was no effect of GS over-expression on photosynthesis or growth. However, under nitrogen starvation the GS transgenics had c. 70% higher shoot and c. 100% greater root dry weight as well as 50% more leaf area than low nitrogen controls. This was achieved by the maintenance of photosynthesis at rates indistinguishable from plants under high nitrogen, while photosynthesis in control plants was inhibited by 40-50% by nitrogen deprivation. It was demonstrated that manipulation of GS activity has the potential to maintain crop photosynthetic productivity while reducing nitrogen fertilization and the concomitant pollution.     

Some effects of nitrate abundance and starvation on metabolism and accumulation of nitrogen in barley (Hordeum vulgare L. cv Sonja)

Nitrate and nitrite reductases were both induced by adding three concentrations of nitrate to the nutrient supply of nitrate-starved barley seedlings. Enzyme induction was not proportional to the amount of nitrate introduced. Glutamine synthetase also increased above a high endogenous activity but the increase did not differ significantly between any of the three nitrate treatments. Nitrate accumulated rapidly in leaves of plants given 4.0 mM or 0.5 mM nitrate but not with 0.1 mM nitrate. In all treatments, amino acids in leaves increased for 2 d, chiefly attributable to glutamine, then declined. Transferring plants from the three nitrate treatments to nitrate-free nutrient produced an immediate decline in nitrate reductase but nitrite reductase continued to increase for 2 d, before declining. Glutamine-synthetase activity was not affected by withdrawal of nitrate, nor did nitrate withdrawal retard plant growth during the 9-d period of the experiment. The disparity between accumulated nitrate and nitrate-reducing capacity and the rapid decrease in leaf nitrate when nutrient nitrate supply was removed, indicated the presence of a nitrate-storage pool that could be called upon to maintain amino-acid production in times of nitrogen starvation.Abbreviations GS glutamine synthetase - NR nitrate reductase - NiR nitrite reductase     

Effects of atrazine on the assimilation of inorganic nitrogen in cereals

The inclusion of sub-lethal amounts ofthe herbicide atrazine [2-chloro-4-(ethylamino)-6-(isopropylamino)-s-triazine] in the nutrient solution supplied to maize and barley increased the growth of the root and shoot and the uptake of nitrate. The activities of nitrate and nitrite reductases, glutamine synthetase and glutamate synthase were enhanced and the amino acid and nitrate contents of the xylem sap increased. All these effects of atrazine were found only in plants grown with nitrate as the nitrogen source. The uptake of ¹⁵NO₃^? and its incorporation into protein in the root and shoot of maize and barley seedlings was significantly greater in the atrazine treated plants. However, a stimulation in the incorporation of leucine-[¹⁴C] into TCA-precipitable protein of detached leaves from 7-day-old barley seedlings was obtained only in the absence of a supply of combined nitrogen either in the culture medium or in the in vitro incubation mixture containing the labelled amino acid.     

大麦氮敏感基因型苗期对氮饥饿的生理响应

以大田试验获得的大麦氮敏感基因型BI-45为材料,利用溶液培养方法,测定了苗期株高、根长、叶绿素含量、含氮量、谷氨酰胺合成酶和硝酸还原酶活性,以及与氮代谢相关的基因（GSI-GSl-2、GSI-3、GS2、Narl、NRT2．J、NRT2-2、NRT2-3和NRT2-4）的表达。结果表明：相对于正常供氮,氮饥饿胁迫下,BI-45根和叶中的氮素利用率提高,含氮量降低,叶绿素含量减少,根冠比增加;叶片中的谷氨酰胺合成酶活性和硝酸还原酶的活性高于根,但是,与叶中的相比,根中的谷氨酰胺合成酶活性升高及硝酸还原酶活性降低的差异性更显著;与正常供氮相比,氮饥饿处理下,根中基因傩家族,基Narl和硝酸盐转运蛋白基因NRT2家族的相对表达量皆达到显著性差异,其中GSl-I、GSl-2和NRT2-2在苗期大麦氮饥饿处理下表现尤为突出,并且在6h都有上调表达。     

The effect of solar UV radiation on four plant species occurring in a coastal grassland vegetation in The Netherlands

During the summer of 1992, growth and some physiological parameters of four native plant species occurring in a coastal grassland in The Netherlands, were studied after reduction of solar UV irradiance using different cut-off filters. Biomass production, morphology and photosynthesis of all species tested were unaffected by the different treatments. Litter production of Plantago lanceolata was increased in the absence of the total UV waveband, indicating a possible role for this waveband in plant senescence. Depletion of the total UV waveband from sunlight resulted in alterations in biomass allocation in Calamagrostis epigeios and Urtica dioica while no changes were observed in P. Ianceolata and Verbascum thapsus. In C. epigeios an increase in the specific leaf area was observed, whereas in U. dioica root weight per total plant weight was decreased resulting in an increase in the shoot/root ratio. Both photosynthetic and UV-absorbing pigment concentrations were altered by the different filter applications. When compared to control plants receiving full sunlight, depletion of UV-B resulted in a significant increase in chlorophyll concentration in U. dioica leaves, this however did not affect photosynthetic rate. The presence of UV-B radiation enhanced the UV-absorbance of leaf extracts of all species except P. lanceolata. Optical characteristics of the leaves were also changed. Both the quantity ( P. lanceolata and U. dioica ) and the quality (all species) of radiation transmitted by the leaves was affected by the different treatments.     

Biochar improved nodulation and nitrogen metabolism of soybean under salt stress

To investigate salt stress and biochar application effects on nodulation and nitrogen metabolism of soybeans (Glycine max cv. M7), an experiment was conducted under the control condition. The treatments comprised three biochar rates (non, 50 and 100 g kg^?1 soil) and three salinities (0, 5 and 10 dS m^?1 NaCl), with four replications of treatments. Salt stress diminished the number of nodules and their weights in the soybean roots. Nitrogen content and metabolism decreased in nodules, roots and shoots, while reducing the activity of glutamate dehydrogenase (GDH), glutamine synthetase (GS), glutamine oxoglutarate aminotransferase (GOGAT) and nitrate reductase (NR). Also, salinity brought down root and shoot weight, total plant biomass, chlorophyll content, leaf area (LA) and rubisco activity in the soybean. On the other hand, application of biochar improved nodulation, nitrogen content, rubisco activity, GDH, GS, GOGAT and NR activities in different parts of the soybean and nodules under salt stress, and consequently improved chlorophyll content, LA, root and shoot weight. Both the 50 and 100 g kg^?1 biochar rates showed similar effects in improving nitrogen metabolism and plant performance under salt stress. Generally, biochar increased nodulation and nitrogen metabolism of the soybean under saline conditions.     

Inorganic nitrogen assimilation in plants of Austrlian rainforest communities

In a study of the plant communities of two Australian rainforests, it was found that pioner species had high levels of nitrate reductase (EC 1.6.6.1) and were predominantly leaf nitrate assimilators. Under- and over-storey species had low levels of shoot and root nitrate reductase activity, and many of them showed little capacity for nitrate reduction even when nitrate ions were freely available. Although closed-forest species have lower levels of nitrate reductase than those of gaps and forest margins, their total nitrogen contents were similar, suggesting the former utilize nitrogen sources other than nitrate ions. Glutamine synthetase (EC 6.3.1.2) was present in the leaves of all species examined. In the leaves of pioneer species the chloroplastic isoform of glutamine synthetase predominted, while in most of the species typical of closed-forest the cytosolic isoform accounted for at least 40% of total leaf activity. Low levels of chloroplastic glutamine synthetase were correlated with a low capacity for leaf nitrate reduction, and both are characteristic of many species that regenerate and grow for some time in shade. Low levels of chloroplastic glutamine synthetase imply that, in some of these woody plants, photorespiratory ammonia is re-assimilated via cytosolic glutamine synthetase.     

The cytosolic glutamine synthetase GLN1;2 plays a role in the control of plant growth and ammonium homeostasis in Arabidopsis rosettes when nitrate supply is not limiting

Glutamine synthetase (EC 6.3.1.2) is a key enzyme of ammonium assimilation and recycling in plants where it catalyses the synthesis of glutamine from ammonium and glutamate. In Arabidopsis, five GLN1 genes encode GS1 isoforms. GLN1;2 is the most highly expressed in leaves and is over-expressed in roots by ammonium supply and in rosettes by ample nitrate supply compared with limiting nitrate supply. It is shown here that the GLN1;2 promoter is mainly active in the minor veins of leaves and flowers and, to a lower extent, in the parenchyma of mature leaves. Cytoimmunochemistry reveals that the GLN1;2 protein is present in the companion cells. The role of GLN1;2 was determined by examining the physiology of gln1;2 knockout mutants. Mutants displayed lower glutamine synthetase activity, higher ammonium concentration, and reduced rosette biomass compared with the wild type (WT) under ample nitrate supply only. No difference between mutant and WT can be detected under limiting nitrate conditions. Despite total amino acid concentration was increased in the old leaves of mutants at high nitrate, no significant difference in nitrogen remobilization can be detected using (15)N tracing. Growing plants in vitro with ammonium or nitrate as the sole nitrogen source allowed us to confirm that GLN1;2 is induced by ammonium in roots and to observe that gln1;2 mutants displayed, under such conditions, longer root hair and smaller rosette phenotypes in ammonium. Altogether the results suggest that GLN1;2 is essential for nitrogen assimilation under ample nitrate supply and for ammonium detoxification.     

Glutamine synthetase of Chlamydomonas: its role in the control of nitrate assimilation

Work is described which suggests that glutamine synthetase (GS) could play an important and direct regulatory role in the control of NO₃ assimilation by the alga. In both steady-state cells and ones disturbed physiologically by changes in light or nitrogen supply the assimilation of NO₃ appears to be limited by the activity of GS. Moreover although in normal cells NH₃ can completely inhibit NO₃ uptake, promote the deactivation of nitrate reductase (NR) and repress the synthesis of NR and nitrite reductase (NIR), these controls are relaxed in cells in which GS is deactivated by treatment with L-methionine-DL-sulfoximine (MSO). It is proposed that the reversible deactivation of GS may play an important part in the regulation of NO₃ assimilation although it is still not clear whether the enzyme itself or products of its metabolism are responsible.Abbreviations GS glutamine synthetase - GS_s glutamine synthetase, synthetase activity - GS_t glutamine synthetase, transferase activity - NR nitrate reductase - NIR nitrite reductase - GDH glutamate dehydrogenase - CHX cycloheximide - MSO L-methionine-DL-sulfoximine - FAD flavine adenine dinucleotide     

Effects of sodium on mineral nutrition in rose plants

The effects of sodium (Na⁺) ion concentration on shoot elongation, uptake of ammonium (NH₄⁺) and nitrate (NO₃^?) and the activities of nitrate reductase (NR) and glutamine synthetase (GS) were studied in rose plants (Rosa hybrida cv. “Lambada”). The results showed that shoot elongation was negatively correlated with sodium concentration, although no external symptoms of toxicity were observed. Nitrate uptake decreased at high sodium levels, specifically at 30 meq litre4 of sodium. As flower development was normal under high saline conditions, this could suggest that nitrogen was being mobilised from shoot and leaf reserves. Ammonium uptake was not affected by any of the salt treatments applied probably because it diffuses through the cell membrane at low concentrations. Nitrate reductase activity was reduced by 50% at 30 meq litre 1 compared with control treatment, probably due to a decrease in the free nitrate related to nitrate uptake pattern. None of the salt treatments used affected total leaf GS activity (both chloroplastic and cytosolic isoforms) or leaf NPK mineral contents. Nitrate reductase activity in leaves increased at 10 meq litre^?1 of sodium and GS activity in roots (cytosolic isoform only) followed the same pattern as NR. It is suggested that the activation of both enzymes at low salt level could be attributed to the beneficial effect of increased sulphur in the nutrient solutions.     

氮素水平对花生氮素代谢及相关酶活性的影响

 在大田高产条件下研究了氮素水平对花生(Arachis hypogaea)可溶性蛋白质、游离氨基酸含量及氮代谢相关酶活性的影响, 结果表明, 适当提高氮素水平既能增加花生各器官中可溶性蛋白质和游离氨基酸的含量, 又能提高硝酸还原酶、谷氨酰胺合成酶和谷氨酸脱氢酶等氮素同化酶的活性, 使其达到同步增加; 氮素水平过高虽能提高硝酸还原酶和籽仁蛋白质含量, 但谷氨酰胺合成酶(GS)和谷氨酸脱氢酶(GDH)的活性下降; N素施肥水平不改变花生植株各器官中可溶性蛋白质、游离氨基酸含量以及硝酸还原酶(NR)、谷氨酰胺合成酶、谷氨酸脱氢酶活性的变化趋势, 但适量施N (A2和A3处理)使花生各营养器官中GS、GDH活性提高; 氮素水平对花生各叶片和籽仁中GS、GDH活性的高低影响较大, 但对茎和根中GDH活性大小的影响较小。     

Evaluation of sole nitrogen sources for shoot and leaf disc cultures of sugarbeet

Use of single nitrogen sources in nutrient media is essential to ascertaining the relative role and regulation of nitrogen assimilatory steps, and may help identify and understand highly productive media for micropropagation and adventitious shoot formation. Eight endogenous nitrogen-containing ions or compounds in sugarbeet (nitrate, ammonium, glutamine, glutamate, urea, proline, glycine betaine and choline) were examined for ability to serve as sole nitrogen source for shoot or leaf disc culture of sugarbeet (Beta vulgaris L.) model clone REL-1. The most productive concentrations of nitrate, ammonium, urea, and glutamine as sole nitrogen sources were moderately supportive of shoot multiplication (64, 70, 81 and 71%, respectively) and fresh weight increase (65, 41, 54 and 41%, respectively) compared to shoot culture growth with the Murashige-Skoog nitrogen mix of 40 mM nitrate and 20 mM ammonium. Glutamate and proline were at best poorly supportive, and glycine betaine and choline were non-supportive. Callus initiation from leaf discs was supported only by nitrate, ammonium, urea, glutamine and proline (50, 100, 100, 100 and 80%, respectively, at the best concentrations, of that on Murashige-Skoog medium). Subsequent shoot regeneration from the intact disc callus in those cultures only occurred on media with nitrate, urea, glutamine, or proline (12, 3, 28 and 3% as many shoots, respectively, as on Murashige-Skoog medium). Overall, the Murashige-Skoog nitrogen mix was superior or equal to any single nitrogen source. However, single nitrogen source media with nitrate, ammonium, urea, glutamine or proline should have significant utility for shoot or leaf disc cultures of mutants with impaired nitrogen assimilation, in comparative physiology studies, or in dual cultures with pathogens of limited ability to use any of these forms of nitrogen. This revised version was published online in June 2006 with corrections to the Cover Date.     

Exogenous proline significantly affects the plant growth and nitrogen assimilation enzymes activities in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis>) under salt stress

Salinity has been shown to be a major factor contributing to low nitrogen availability in plants. To verify the changes in nitrogen metabolism activity as affected by the exogenous application of proline under salt stress and its relation to salt tolerance, in vitro rice shoot apices were used as a model to study the growth performance and changes in nitrogen assimilation activities in two Malaysian rice cultivars MR 220 and MR 253. Results revealed that salt stress greatly reduced the plant height, shoot nitrate (NO₃ ^?) content, shoot glutamine synthetase (GS), and root nitrate reductase (NR) activities in both cultivars. Supplementation of proline significantly increased the plant height, number of roots, root NO₃ ^? content, root NR, and root GS activities under salt stress in both cultivars with greater enhancement in MR 253 than MR 220. The results also indicated that MR 253 possessed higher nitrite reductase (NiR) and glutamate synthase (NADH–GOGAT) activities as compared with MR 220 in all tested treatments. It was suggested that the NO₃ ^? content, NR, and GS activities played important roles in regulating nitrogen metabolism under salt stress. Taken together, it was concluded that the ability of proline in mitigating salt stress-induced damages was correlated with the changes in nitrogen assimilation activities.     

Appearance of nitrate reductase, nitrite reductase and glutamine synthetase in different organs of the Scots pine (Pinus sylvestris) seedling as affected by light, nitrate and ammonium

Appearance of nitrate reductase (NR, EC 1.6.6.1–3), nitrite reductase (NiR, EC 1.7.7.1) and glutamine synthetase (GS, EC 6.3.1.2) under the control of nitrate, ammonium and light was studied in roots, hypocotyls and needles (cotyledonary whorl) of the Scots pine ( Pinus sylvestris L.) seedling. It was found that appearance of NiR was mainly controlled by nitrate whereas appearance of GS was strongly controlled by light. In principle, the NR activity level showed the same dependency on nitrate and light as that of NiR. In the root, both nitrate and ammonium had a stimulatory effect on GS activity whereas in the whorl the induction was minor. The level of NiR (NR) activity is high in the root and hypocotyl and low in the cotyledonary whorl, whereas the GS activity level per organ increases strongly from the root to the whorl. Thus, in any particular organ the operation of the glutamine synthetase/glutamate synthase (GS/GOGAT) cycle is not closely connected to the operation of the nitrate reduction pathway. The strong control of GS/GOGAT by light and the minor sensitivity to induction by nitrate or ammonium indicate a major role of the GS/GOGAT cycle in reassimilation of endogeniously generated ammonium.     

Nitrogen metabolism of Plantago major ssp. major as dependent on the supply of mineral nutrients

Various aspects of the nitrogen metabolism of Plantago major L. ssp. major , a grassland species from a relatively nutrient-rich habitat, were investigated with plants that were grown in nutrient-rich and nutrient-poor culture solutions. In addition the influence of a change in the nutrient supply, both from high to low and from low to high nutrient conditions was studied. The data on P. major major showed that the activities of nitrate reductase (NR) and the reduced nitrogen content were all rather independent of the nutrient supply. These findings are in contrast with those of P. lanceolata , a grassland species from a relatively nutrient-poor habitat, where both the activities of NR, GDH and GS and the reduced nitrogen content were correlated with the level of the nutrient supply. The rigidity of the nitrogen metabolism of P. major major may be of advantage if the nutrient supply is limited only for relatively short periods.     

Effect of ammonium salt on enzymes of ammonium assimilation in maize seedlings

Glutamate dehydrogenase (GDH E.C. 1.4.1.2.4), glutamine synthetase (GS E.C. 6.3.1.2) and glutamate synthase (glutamine oxoglutarate amino transferase, GOGAT E.C. 2.6.1.53) activities, protein and organic nitrogen contents and growth of roots and shoots of maize seedlings raised in dark at 25±2°C in half strength Hoagland’s solution containing different ammonium salts as source of nitrogen, were determined to assess the contribution of alternate pathways in ammonium assimilation. Ammonium nitrate or in some cases ammonium chloride appeared to be the best source for both root and shoot growth and for increase in protein, total nitrogen and the enzymes of ammonium assimilation. In roots, NH₄-nitrogen appeared to be assimilated by both GDH as well as GS-GOGAT pathways specially in the dark grown seedlings, while in shoots it was primarily by GS-GOGAT pathway.     

Tissue and Cellular Distribution of Glutamine Synthetase in Roots of Pea (Pisum sativum) Seedlings

The effect of nitrate application on glutamine synthetase activity in roots of pea (Pisum sativum L.) seedlings (2 weeks old) was studied. Separation of organelles from root fragments by sucrose density-gradient centrifugation revealed that both nitrite reductase and glutamine synthetase activities increased in root plastids as a response to nitrate application and that no such response was induced by ammonium application. Glutamine synthetase activity was also found to increase in plastids with distance from apex in nitrate-treated plants, the highest specific activity being located in the fourth 1-centimeter segment. Separation by SDS-PAGE and characterization by Western blotting showed that cytosolic glutamine synthetase contains one subunit polypeptide (28 kilodaltons) and that plastid glutamine synthetase contains both the 38-kilodalton subunit and a heavier subunit. When nitrate was present in the nutrient solution, the heavier subunit increased in abundance in protein fractions obtained from purified root plastids.     

Growth and nitrogen metabolism of Catasetum fimbriatum (orchidaceae) grown with different nitrogen sources

Catasetum fimbriatum is an epiphytic orchid from South America that has been used for 15 years as a model plant for metabolic and developmental studies in our laboratory. In this work, C. fimbriatum plants were aseptically grown with 6 mol m(-3) of either glutamine or inorganic nitrogen forms (NO(3)(-):NH(4)(+) ratios). The highest biomass accumulation was found in plants supplied with glutamine; no significant difference was observed in plants incubated in the presence of inorganic nitrogen sources. Nitrogen assimilation was limited in the presence NO(3)(-) as a sole nitrogen source. C. fimbriatum did not accumulate NO(3)(-) and very low rates of in vivo nitrate reductase activity were observed. Most nitrate reductase activity (70%) was detected in the 2 cm apical roots. Nitrate-treated plants exhibited relatively lower amounts of free amino-N, chlorophyll and free NH(4)(+) contents and higher soluble sugar contents than the NH(4)(+)-treated plants. While shoot glutamine synthetase activity was only slightly affected by nitrogen sources, root glutamine synthetase activity was not modified by any nitrogen form. Glutamate dehydrogenase-NADH activity in shoot tissues was not influenced by any nitrogen source. However, the glutamate dehydrogenase-NADH activity in roots was enhanced when NH(4)(+) tissue contents was augmented by increasing NH(4)(+) in the medium and by the presence of glutamine. Our results strongly suggest that organic nitrogen and NH(4)(+) are probably the most important nitrogen sources to C. fimbriatum plants.     

Carbon and nitrogen metabolism in the seagrass, Zostera marina L.: Environmental control of enzymes involved in carbon allocation and nitrogen assimilation

This study experimentally examined influences of environmental variables on the activities of key enzymes involved in carbon and nitrogen metabolism of the submersed marine angiosperm, Zostera marina L. Nitrate reductase activity in leaf tissue was correlated with both water-column nitrate concentrations and leaf sucrose levels. Under elevated nitrate, shoot nitrate reductase activity increased in both light and dark periods if carbohydrate reserves were available. When water-column nitrate was low, glutamine synthetase activity in leaf tissue increased with environmental ammonium. In contrast, glutamine synthetase activity in belowground tissues was statistically related to both nitrate and temperature. At the optimal growth temperature for this species (ca. 25 °C), increased water-column nitrate promoted an increase in glutamine synthetase activity of belowground tissues. As temperatures diverged from the optimum, this nitrate effect on glutamine synthetase was no longer evident. Activities of both sucrose synthase and sucrose-P synthase were directly correlated with temperature. Sucrose-P synthase activity also was correlated with salinity, and sucrose synthase activity was statistically related to tissue ammonium. Overall, the enzymatic responses that were observed indicate a tight coupling between carbon and nitrogen metabolism that is strongly influenced by prevailing environmental conditions, especially temperature, salinity, and environmental nutrient levels.