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1.
Nitrate reductase activity in excised embryos of Agrostemma githago increases in response to both NO 3− and cytokinins. We asked the question whether cytokinins affected nitrate reductase activity directly or through NO 3−, either by amplifying the effect of low endogenous NO 3− levels, or by making NO 3− available for induction from a metabolically inactive compartment. Nitrate reductase activity was enhanced on the average by 50% after 1 hour of benzyladenine treatment. In some experiments, the cytokinin response was detectable as early as 30 minutes after addition of benzyladenine. Nitrate reductase activity increased linearly for 4 hours and began to decay 13 hours after start of the hormone treatment. When embryos were incubated in solutions containing mixtures of NO 3− and benzyladenine, additive responses were obtained. The effects of NO 3− and benzyladenine were counteracted by abscisic acid. The increase in nitrate reductase activity was inhibited at lower abscisic acid concentrations in embryos which were induced with NO 3−, as compared to embryos treated with benzyladenine. Casein hydrolysate inhibited the development of nitrate reductase activity. The response to NO 3− was more susceptible to inhibition by casein hydrolysate than the response to the hormone. When NO 3− and benzyladenine were withdrawn from the medium after maximal enhancement of nitrate reductase activity, the level of the enzyme decreased rapidly. Nitrate reductase activity increasd again as a result of a second treatment with benzyladenine but not with NO 3−. At the time of the second exposure to benzyladenine, no NO 3− was detectable in extracts of Agrostemma embryos. This is taken as evidence that cytokinins enhance nitrate reductase activity directly and not through induction by NO 3−. 相似文献
2.
The effect of nitrogen form (NH 4-N, NH 4-N + NO 3−, NO 3−) on nitrate reductase activity in roots and shoots of maize ( Zea mays L. cv INRA 508) seedlings was studied. Nitrate reductase activity in leaves was consistent with the well known fact that NO 3− increases, and NH 4+ and amide-N decrease, nitrate reductase activity. Nitrate reductase activity in the roots, however, could not be explained by the root content of NO 3−, NH 4-N, and amide-N. In roots, nitrate reductase activity in vitro was correlated with the rate of nitrate reduction in vivo. Inasmuch as nitrate reduction results in the production of OH − and stimulates the synthesis of organic anions, it was postulated that nitrate reductase activity of roots is stimulated by the released OH − or by the synthesized organic anions rather than by nitrate itself. Addition of HCO 3− to nutrient solution of maize seedlings resulted in a significant increase of the nitrate reductase activity in the roots. As HCO 3−, like OH −, increases pH and promotes the synthesis of organic anions, this provides circumstantial evidence that alkaline conditions and/or organic anions have a more direct impact on nitrate reductase activity than do NO 3−, NH 4-N, and amide-N. 相似文献
3.
Growth chamber studies with soybeans ( Glycine max [L.] Merr.) were designed to determine the relative limitations of NO 3−, NADH, and nitrate reductase (NR) per se on nitrate metabolism as affected by light and temperature. Three NR enzyme assays (+NO 3−in vivo, −NO 3−in vivo, and in vitro) were compared. NR activity decreased with all assays when plants were exposed to dark. Addition of NO 3− to the in vivo NR assay medium increased activity (over that of the −NO 3−in vivo assay) at all sampling periods of a normal day-night sequence (14 hr-30 C day; 10 hr-20 C night), indicating that NO 3− was rate-limiting. The stimulation of in vivo NR activity by NO 3− was not seen in plants exposed to extended dark periods at elevated temperatures (16 hr-30 C), indicating that under those conditions, NO 3− was not the limiting factor. Under the latter condition, in vitro NR activity was appreciable (19 μmol NO 2− [g fresh weight, hr] −1) suggesting that enzyme level per se was not the limiting factor and that reductant energy might be limiting. 相似文献
4.
Selected variant cell lines of Haplopappus gracilis (Nutt) Gray that showed disturbed growth after transfer from an alanine medium to NO 3− medium were characterized. The in vivo NO 3− reductase activity (NRA) was lower in these lines than in the wild type. In vitro NRA assays suggest that decreased in vivo NRA was not caused by a lower amount of active enzyme. Cells of the variant lines revealed up to 75% lower extractable activity of NO 2− reductase as compared with the wild type. This coincided with higher accumulation of NO 2− by the variant than by the wild type cells after transfer from alanine medium to NO 3− medium. NO 2− accumulation was transient or continuous, depending on cell line, metabolic state of the cells, and light conditions. 相似文献
5.
Coastal zones act as a sink for riverine and atmospheric nitrogen inputs and thereby buffer the open ocean from the effects of anthropogenic activity. Recently, microbial activity in sandy permeable sediments has been identified as a dominant source of N-loss in coastal zones, namely through denitrification. Some of the highest coastal denitrification rates measured so far occur within the intertidal permeable sediments of the eutrophied Wadden Sea. Still, denitrification alone can often account for only half of the substantial nitrate (NO 3
−) consumption. Therefore, to investigate alternative NO 3
− sinks such as dissimilatory nitrate reduction to ammonium (DNRA), intracellular nitrate storage by eukaryotes and isotope equilibration effects we carried out 15NO 3
− amendment experiments. By considering all of these sinks in combination, we could quantify the fate of the 15NO 3
− added to the sediment. Denitrification was the dominant nitrate sink (50–75%), while DNRA, which recycles N to the environment accounted for 10–20% of NO 3
− consumption. Intriguingly, we also observed that between 20 and 40% of 15NO 3
− added to the incubations entered an intracellular pool of NO 3
− and was subsequently respired when nitrate became limiting. Eukaryotes were responsible for a large proportion of intracellular nitrate storage, and it could be shown through inhibition experiments that at least a third of the stored nitrate was subsequently also respired by eukaryotes. The environmental significance of the intracellular nitrate pool was confirmed by in situ measurements which revealed that intracellular storage can accumulate nitrate at concentrations six fold higher than the surrounding porewater. This intracellular pool is so far not considered when modeling N-loss from intertidal permeable sediments; however it can act as a reservoir for nitrate during low tide. Consequently, nitrate respiration supported by intracellular nitrate storage can add an additional 20% to previous nitrate reduction estimates in intertidal sediments, further increasing their contribution to N-loss. 相似文献
6.
Soybean ( Glycine max [L.] Merr.) seeds were imbibed and germinated with or without NO 3−, tungstate, and norflurazon (San 9789). Norflurazon is a herbicide which causes photobleaching of chlorophyll by inhibiting carotenoid synthesis and which impairs normal chloroplast development. After 3 days in the dark, seedlings were placed in white light to induce extractable nitrate reductase activity. The induction of maximal nitrate reductase activity in greening cotyledons did not require NO 3− and was not inhibited by tungstate. Induction of nitrate reductase activity in norflurazon-treated cotyledons had an absolute requirement for NO 3− and was completely inhibited by tungstate. Nitrate was not detected in seeds or seedlings which had not been treated with NO 3−. The optimum pH for cotyledon nitrate reductase activity from norflurazon-treated seedlings was at pH 7.5, and near that for root nitrate reductase activity, whereas the optimum pH for nitrate reductase activity from greening cotyledons was pH 6.5. Induction of root nitrate reductase activity was also inhibited by tungstate and was dependent on the presence of NO 3−, further indicating that the isoform of nitrate reductase induced in norflurazon-treated cotyledons is the same or similar to that found in roots. Nitrate reductases with and without a NO 3− requirement for light induction appear to be present in developing leaves. In vivo kinetics (light induction and dark decay rates) and in vitro kinetics (Arrhenius energies of activation and NADH:NADPH specificities) of nitrate reductases with and without a NO 3− requirement for induction were quite different. Km values for NO 3− were identical for both nitrate reductases. 相似文献
7.
Previously, we reported that in Citrus plants, nitrate influx through the plasmalemma of roots cells follows a biphasic pattern, suggesting the existence of at least two different uptake systems, a high and low affinity transport system (HATS and LATS, respectively). Here, we describe a novel inducible high affinity transport system (iHATS). This new nitrate transport system has a high capacity to uptake nitrate in two different Citrus rootstocks (Cleopatra mandarin and Troyer citrange). The iHATS was saturable, showing higher affinity than constitutive high affinity transport system (cHATS) to the substrate NO 3−. The V max for this saturable component iHATS was higher than cHATS, reaching similar values in both rootstocks.Additionally, we studied the regulation of root NO 3− uptake mediated by both HATS (iHATS and cHATS) and LATS. In both rootstocks, cHATS is constitutive and independent of N-status. Concerning the regulation of iHATS, this system is upregulated by NO 3− and down-regulated by the N status and by NO 3− itself when plants are exposed to it for a longer period of time. LATS in Cleopatra mandarin and Troyer citrange rootstocks is repressed by the N-status.The use of various metabolic uncouplers or inhibitors indicated that NO 3− net uptake mediated by iHATS and LATS was an active transport system in both rootstocks.Key Words: Citrus, inducible high affinity transport system (iHATS), constitutive high affinity transport system (cHATS), nitrate uptake, regulation 相似文献
8.
The objective of this study was to identify factors which limit leaf nitrate reductase (NR) activity as decline occurs during flowering and beginning seed development in soybean ( Glycine max [L.] Merr. cv Clark). Level of NR enzyme activity, level of reductant, and availability of NO 3− as substrate were evaluated for field-grown soybean from flowering through leaf senescence. Timing of reproductive development was altered within one genotype by (a) exposure of Clark to an artificially short photoperiod to hasten flowering and podfill, and (b) the use of an early flowering isoline. Nitrogen (N) was soil-applied to selected plots at 500 kilograms per hectare as an additional variable. Stem NO 3− concentration and in vivo leaf NR activity were significantly correlated ( R2 = 0.69 with nitrate in the assay medium and 0.74 without nitrate in the medium at P = 0.001) across six combinations of reproductive and soil N-treatment. The supply of NO 3− from the root to the leaf tissue was the primary limitation to leaf NR activity during flowering and podfill. Levels of NR enzyme and reductant were not limiting to leaf NR activity during this period. 相似文献
9.
The comparative induction of nitrate reductase (NR) by ambient NO 3− and NO 2− as a function of influx, reduction (as NR was induced) and accumulation in detached leaves of 8-day-old barley ( Hordeum valgare L.) seedlings was determined. The dynamic interaction of NO 3− influx, reduction and accumulation on NR induction was shown. The activity of NR, as it was induced, influenced its further induction by affecting the internal concentration of NO 3−. As the ambient concentration of NO 3− increased, the relative influences imposed by influx and reduction on NO 3− accumulation changed with influx becoming a more predominant regulant. Significant levels of NO 3− accumulated in NO 2−-fed leaves. When the leaves were supplied cycloheximide or tungstate along with NO 2−, about 60% more NO 3− accumulated in the leaves than in the absence of the inhibitors. In NO 3−-supplied leaves NR induction was observed at an ambient concentration of as low as 0.02 m m. No NR induction occurred in leaves supplied with NO 2− until the ambient NO 2− concentration was 0.5 m m. In fact, NR induction from NO 2− solutions was not seen until NO 3− was detected in the leaves. The amount of NO 3− accumulating in NO 2−-fed leaves induced similar levels of NR as did equivalent amounts of NO 3− accumulating from NO 3−-fed leaves. In all cases the internal concentration of NO 3−, but not NO 2−, was highly correlated with the amount of NR induced. The evidence indicated that NO 3− was a more likely inducer of NR than was NO 2−. 相似文献
10.
Chlamydomonas reinhardii cells, growing photoautotrophically under air, excreted to the culture medium much higher amounts of NO 2− and NH 4+ under blue than under red light. Under similar conditions, but with NO 2− as the only nitrogen source, the cells consumed NO 2− and excreted NH 4+ at similar rates under blue and red light. In the presence of NO 3− and air with 2% CO 2 (v/v), no excretion of NO 2− and NH 4+ occurred and, moreover, if the bubbling air of the cells that were currently excreting NO 2− and NH 4+ was enriched with 2% CO 2 (v/v), the previously excreted reduced nitrogen ions were rapidly reassimilated. The levels of total nitrate reductase and active nitrate reductase increased several times in the blue-light-irradiated cells growing on NO 3− under air. When tungstate replaced molybdate in the medium (conditions that do not allow the formation of functional nitrate reductase), blue light activated most of the preformed inactive enzyme of the cells. Furthermore, nitrate reductase extracted from the cells in its inactive form was readily activated in vitro by blue light. It appears that under high irradiance (90 w m −2) and low CO 2 tensions, cells growing on NO 3− or NO 2− may not have sufficient carbon skeletons to incorporate all the photogenerated NH 4+. Because these cells should have high levels of reducing power, they might use NO 3− or, in its absence, NO 2− as terminal electron acceptors. The excretion of the products of NO 2− and NH 4+ to the medium may provide a mechanism to control reductant level in the cells. Blue light is suggested as an important regulatory factor of this photorespiratory consumption of NO 3− and possibly of the whole nitrogen metabolism in green algae. 相似文献
11.
The effect of water stress on patterns of nitrate reductase activity in the leaves and nodules and on nitrogen fixation were investigated in Medicago sativa L. plants watered 1 week before drought with or without NO 3−. Nitrogen fixation was decreased by water stress and also inhibited strongly by the presence of NO 3−. During drought, leaf nitrate reductase activity (NRA) decreased significantly particularly in plants watered with NO 3−, while with rewatering, leaf NRA recovery was quite important especially in the NO 3−-watered plants. As water stress progressed, the nodular NRA increased both in plants watered with NO 3− and in those without NO 3− contrary to the behavior of the leaves. Beyond −15.10 5 pascal, nodular NRA began to decrease in plants watered with NO 3−. This phenomenon was not observed in nodules of plants given water only. 相似文献
12.
Up to 80% of the total nitrate reductase activity (NRA) determined in vivo in different parts of vegetative tobacco plant ( Nicotiana tabacum) was located in the leaves. The NRA reached a peak when a leaf had expanded to 27% of its final weight and 33% of its final area. Thereafter, with advancing expansion and age of the leaf, the activity declined. This pattern of development of NRA during the ontogenesis of leaves was not influenced by raising the supply of NO 3− from 3 to 6 milliequivalent per cubic decimeter in the substrate solution. The concentration of NO 3− in leaves, stem and root was inversely related to NRA at both NO 3− levels. Raising the supply of K + from 1 to 6 milliequivalent per cubic decimeter at either concentration of NO 3− slowed down the development of NRA in the initial stages of expansion, but promoted it subsequently. The peak of the activity which developed in a leaf of 62% of its final area was higher at the higher supply of K +. The higher activity was maintained thereafter in the expanding and in matured and older leaves. It was concluded that NRA and the pattern of its development in expanding leaves is related to the availability of metabolites and their incorporation into enzyme proteins. Both these processes are influenced by: (a) the vertical profile of concentration of K + in the shoot and (b) the concentration of K + in a leaf, which depend upon its supply. 相似文献
13.
The photoreversible nature of the regulation of nitrate reductase is one of the most interesting features of this enzyme. As well as other chemicals, NH 2OH reversibly inactivates the reduced form of nitrate reductase from Ankistrodesmus braunii. From the partial activities of the enzyme, only terminal nitrate reductase is affected by NH 2OH. To demonstrate that the terminal activity was readily inactivted by NH 2OH, the necessary reductants of the terminal part of the enzyme had to be cleared of dithionite since this compound reacts chemically with NH 2OH. Photoreduced flavins and electrochemically reduced methyl viologen sustain very effective inactivation of terminal nitrate reductase activity, even if the enzyme was previously deprived of its NADH-dehydrogenase activity. The early inhibition of nitrate reductase by NH 2OH appears to be competitive versus NO 3−. Since NO 3−, as well as cyanate, carbamyl phosphate and azide (competitive inhibitors of nitrate reductase versus NO 3−), protect the enzyme from NH 2OH inactivation, it is suggested that NH 2OH binds to the nitrate active site. The NH 2OH-inactivated enzyme was photoreactivated in the presence of flavins, although slower than when the enzyme was previously inactivated with CN −. NH 2OH and NADH concentrations required for full inactivation of nitrate reductase appear to be low enough to potentially consider this inactivation process of physiological significance. 相似文献
14.
The cytoplasmic NO 3− concentration ([NO 3−] c) was estimated for roots of barley ( Hordeum vulgare L. cv Klondike) using a technique based on measurement of in vivo nitrate reductase activity. At zero external NO 3− concentration ([NO 3−] o), [NO 3−] c was estimated to be 0.66 m m for plants previously grown in 100 μ m NO 3−. It increased linearly with [NO 3−] o between 2 and 20 m m, up to 3.9 m m at 20 m m [NO 3−] o. The values obtained are much lower than previous estimates from compartmental analysis of barley roots. These observations support the suggestion (MY Siddiqi, ADM Glass, TJ Ruth [1991] J Exp Bot 42: 1455-1463) that the nitrate reductase-based technique and compartmental analysis determine [NO 3−] c for two separate pools; an active, nitrate reductase-containing pool (possibly located in the epidermal cells) and a larger, slowly metabolized storage pool (possibly in the cortical cells), respectively. Given the values obtained for [NO 3−] c and cell membrane potentials of −200 to −300 mV (ADM Glass, JE Schaff, LV Kochian [1992] Plant Physiol 99: 456-463), it is very unlikely that passive influx of NO 3− is possible via the high-concentration, low-affinity transport system for NO 3−. This conclusion is consistent with the suggestion by Glass et al. that this system is thermodynamically active and capable of transporting NO 3− against its electrochemical potential gradient. 相似文献
15.
An experiment was conducted to test the hypothesis that, when nitrogenase and nitrate reductase both contribute to the nitrogen nutrition of a nodulated legume, nitrogenase activity is inversely proportional to the rate of accumulation of organic nitrogen derived from the reduction of nitrate. Trifolium subterraneum L. plants, inoculated with Rhizobium trifolii and sown as small swards, were allowed to establish a closed canopy and steady rates of growth, dinitrogen fixation, and nitrogen accumulation. Swards were then supplied with nutrient solutions of 0, 0.5, 1.0, or 2.5 m m NO 3− with a 29.69% enrichment of 15N and allowed to grow for a further 33 days. Harvests were made to measure dry weight, nitrogen accumulation, 15N accumulation, NO 3− content and nitrogenase activity by acetylene reduction assay. Since the 15N of the plant organic matter could have been derived only from the NO 3− of the nutrient solution, its rate of accumulation provided a measure of the rate of NO 3− reduction. It was found that as this rate increased in response to external NO 3− concentration the rate of nitrogenase activity decreased proportionately. It is concluded that the reduction of nitrate and the reduction of dinitrogen act in a complementary manner to supply a plant with organic nitrogen for growth. 相似文献
16.
The effects of CO 2-limited photosynthesis on 15NO 3− uptake and reduction by maize ( Zea mays, DeKalb XL-45) seedlings were examined in relation to concurrent effects of CO 2 stress on carbohydrate levels and in vitro nitrate reductase activities. During a 10-hour period in CO 2-depleted air (30 microliters of CO 2/ per liter), cumulative 15NO 3− uptake and reduction were restricted 22 and 82%, respectively, relative to control seedlings exposed to ambient air containing 450 microliters of CO 2 per liter. The comparable values for roots of decapitated maize seedlings, the shoots of which had previously been subjected to CO 2 stress, were 30 and 42%. The results demonstrate that reduction of entering nitrate by roots as well as shoots was regulated by concurrent photosynthesis. Although in vitro nitrate reductase activity of both tissues declined by 60% during a 10-hour period of CO 2 stress, the remaining activity was greatly in excess of that required to catalyze the measured rate of 15NO 3− reduction. Root respiration and soluble carbohydrate levels in root tissue were also decreased by CO 2 stress. Collectively, the results indicate that nitrate uptake and reduction were regulated by the supply of energy and carbon skeletons required to support these processes, rather than by the potential enzymatic capacity to catalyze nitrate reduction, as measured by in vitro nitrate reductase activity. 相似文献
17.
Studies were conducted to quantitate the evolution of nitrogen oxides (NO (x)) from soybean [ Glycine max (L.) Merr.] leaves during in vivo nitrate reductase (NR) assays with aerobic and anaerobic gas purging. Anaerobic gas purging (N 2 and argon) consistently resulted in greater NO (x) evolution than did aerobic gas purging (air and O 2). The evolution of NO (x) was dependent on gas flow rate and on NO 2− formation in the assay medium; although a threshold level of NO 2− appeared to exist beyond which the rate of NO (x) evolution did not increase further. 相似文献
18.
Effects of Na application on the capacity of NO 3− assimilation were studied in Na-deficient Amaranthus tricolor L. cv Tricolor plants. On day 30 after germination, Na-deficient A. tricolor plants received either 0.5 millimolar NaCl or KCl. The level of nitrate reductase activity doubled within 24 hours by the addition of Na and the enhanced level was maintained thereafter. When the plants were exposed to 2 millimolar 15NO 3−, total 15N taken up by the plants was greater in the Na-treated plants than in the K-treated plants within 24 hours of the Na treatment. Incorporation of 15N into the 80% ethanol-insoluble nitrogen fraction of the Na-treated plants in the light period was about 260% of those of the K-treated plants indicating greater capacity of NO 3− assimilation in the Na-treated plants. From these results, it was demonstrated that Na application to the Na-deficient A. tricolor plants promoted NO 3− reduction and its subsequent assimilation into protein, resulting in growth enhancement. 相似文献
19.
Ricinus communis L. plants were grown in nutrient solutions in which N was supplied as NO 3− or NH 4+, the solutions being maintained at pH 5.5. In NO 3−-fed plants excess nutrient anion over cation uptake was equivalent to net OH − efflux, and the total charge from NO 3− and SO 42− reduction equated to the sum of organic anion accumulation plus net OH − efflux. In NH 4+-fed plants a large H + efflux was recorded in close agreement with excess cation over anion uptake. This H + efflux equated to the sum of net cation (NH 4+ minus SO 42−) assimilation plus organic anion accumulation. In vivo nitrate reductase assays revealed that the roots may have the capacity to reduce just under half of the total NO 3− that is taken up and reduced in NO 3−-fed plants. Organic anion concentration in these plants was much higher in the shoots than in the roots. In NH 4+-fed plants absorbed NH 4+ was almost exclusively assimilated in the roots. These plants were considerably lower in organic anions than NO 3−-fed plants, but had equal concentrations in shoots and roots. Xylem and phloem saps were collected from plants exposed to both N sources and analyzed for all major contributing ionic and nitrogenous compounds. The results obtained were used to assist in interpreting the ion uptake, assimilation, and accumulation data in terms of shoot/root pH regulation and cycling of nutrients. 相似文献
20.
The effect of NaCl and Na 2SO 4 salinity on NO 3− assimilation in young barley ( Hordeum vulgare L. var Numar) seedlings was studied. The induction of the NO 3− transporter was affected very little; the major effect of the salts was on its activity. Both Cl − and SO 42− salts severely inhibited uptake of NO 3−. When compared on the basis of osmolality of the uptake solutions, Cl − salts were more inhibitory (15-30%) than SO 42− salts. At equal concentrations, SO 42− salts inhibited NO 3− uptake 30 to 40% more than did Cl − salts. The absolute concentrations of each ion seemed more important as inhibitors of NO 3− uptake than did the osmolality of the uptake solutions. Both K + and Na + salts inhibited NO 3− uptake similarly; hence, the process seemed more sensitive to anionic salinity than to cationic salinity. Unlike NO3− uptake, NO3− reduction was not affected by salinity in short-term studies (12 hours). The rate of reduction of endogenous NO3− in leaves of seedlings grown on NaCl for 8 days decreased only 25%. Nitrate reductase activity in the salt-treated leaves also decreased 20% but its activity, determined either in vitro or by the `anaerobic' in vivo assay, was always greater than the actual in situ rate of NO3− reduction. When salts were added to the assay medium, the in vitro enzymic activity was severely inhibited; whereas the anaerobic in vivo nitrate reductase activity was affected only slightly. These results indicate that in situ nitrate reductase activity is protected from salt injury. The susceptibility to injury of the NO3− transporter, rather than that of the NO3− reduction system, may be a critical factor to plant survival during salt stress. 相似文献
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