Devenir de la putrescine-1,4-14C chez Glycine max

Fate of Putrescine-1,4-¹⁴C in Glycine max Putrescine-1,4-¹⁴C was supplied to young decotylized Glycine max (L.) Merr. cv. Chippewa plants growing under aseptic conditions on a liquid medium with nitrogen supplied either as ammonium chloride or nitrates. Whatever the source of nitrogen the diamine was quickly transformed to γ-aminobutyric acid, succinic acid and malic acid; only a very minor part was utilized for the synthesis of polyamines. In the presence of ammonium chloride the putrescine catabolism may be slower than in the presence of nitrates. The results are explained by a weak isotopic dilution or by a diamine oxidase activity lower in “ammonium” plants than in “nitrate” plants; the two causes might co-exist. The possibility of a diamine compartmentation has to be considered.     

Etude de l'activite de l'arginine decarboxylase dans les jeunes plantes de Glycine max privees de leurs cotyledons

l-Arginine carboxy-lyase and l-ornithine carboxy-lyase activities were investigated in young decotylized Glycine max plants growing in the light on a liquid medium in the presence of ammonium chloride or nitrate. Only the first enzyme could be detected. It had pH optimum at 7.0 activity and there is much more activity in plants cultivated on ammonium chloride than in plants grown on nitrate.     

The effects of nitrate on the enzymes involved in nitrogen assimilation in nodulated pea roots

Pea Plants ( Pisum sativaum L. ev. Little Marvel) were grown in N-free medium and when well nodulated (28 days) were supplied for 8 days with nitrate or ammonium. Over the 8 days of nitrate treatment, total amino and amide N in sap declined, and the proportion of aspartate relative to the other amino acids increased. After 8 days of treatment, nitrogenase (EC 1.18.2.1) activity in nitrate-treated plants declined to about 30% of the activity in controls even though nodules were not directly in contact with nutrient solution. Nitrogenase activity was also decreased by the addition of ammonium chloride (10 m M ). With addition of nitrate or ammonium. clear signs of senescence began to show in the nodules after 4 days. Nitrate reductase (EC 1.6.6.1) activity was induced in roots by nitrate, but decreased sharply in nodules. In response to nitrate addition, newly formed root tissues showed 3- to 5-times higher glutamine synthetase (GS. EC 6.3.1.4) activity than newly formed tissues of control plants, expressed on a protein or weight basis. In complementary experiments, when ammonium salts were used instead of nitrates, the increase in GS activity was significantly lower. GS activity decreased in nodules of treated plants and total extractable protein was 3 times lower in nodules of nitrate-treated plants than in controls at day 8 of treatment.     

Cytokinin-dependent expression of the ARR5::GUS construct during transgenic arabidopsis growth

Growth and glucuronidase (GUS) activity were followed in the cotyledons and rosette leaves of Arabidopsis thaliana (L.) Heynh (ecotype Wassilewskija) plants transformed with the GUS gene under the control of the cytokinin-dependent promoter of the ARR5 gene. The presence of active cytokinins in plant tissues was assessed from GUS activity. Plants were grown for three weeks on the nitrate-or ammonium-containing nutrient medium. In plants grown on ammonium nutrition, cotyledon and leaf growth was substantially suppressed as compared with plants feeding with nitrates. In correspondence with this growth inhibition, GUS activity was markedly lower in plant leaves grown on the ammonium-containing medium. This indicated a reduction in these leaves of active cytokinin forms capable of activation of the promoter for the ARR5 gene. On both nitrogen sources, GUS activity increased during leaf growth and dropped sharply after growth ceasing. This indicated that leaf growth depended on the cytokinin content in them. High GUS activity was detected in petioles and leaf conductive system, indicating leaf providing with cytokinins along the conductive vessels. A sharp drop in the GUS activity after leaf growth stoppage coincided in time with GUS activation in the leaf positioned above this leaf. This indicated possible cytokinin redistribution in the plant; its content could be a limiting factor for leaf growth. A higher growth rate in plants on nitrate nitrogen nutrition and corresponding high GUS activity in them are discussed in terms of cytokinin signaling role in leaf growth regulation mediated by nitrate.     

Effect of Transpiration-reducing Chemicals on Growth, Flowering, and Stomatal Opening of Tomato Plants

Phenyl mercuric acetate, 8-hydroxyquinoline, N-dimethylamino succinamic acid, or 2-chloroethyl trimethyl ammonium chloride were sprayed on 37-day-old tomato (Lycopersicon esculentum Mill. cv. Sioux) plants seven times at weekly intervals. Plants of nearly normal appearance resulted with all treatments except 2-chloroethyl trimethyl ammonium chloride. There was no change in leaf number, but 2-chloroethyl trimethyl ammonium chloride increased the number of flowers. 2-Chloroethyl trimethyl ammonium chloride and phenyl mercuric acetate caused earlier flowering. Yield was not affected significantly. Stomatal opening was reduced 80% immediately after spraying with phenyl mercuric acetate or 2-chloroethyl trimethyl ammonium chloride, but 6 days after spraying, the reduction in stomatal opening was only 30 to 40%. Wilting was delayed 8 days by phenyl mercuric acetate and 4 days by 2-chloroethyl trimethyl ammonium chloride and N-dimethyl amino succinamic acid treatments, when water was withheld 59 days after the final spray application.     

Salt tolerance in the halophyte Suaeda maritima. Further properties of the enzyme malate dehydrogenase

Malate dehydrogenase activity in supernatant fractions prepared from the halophyte Suaeda maritima was modified by added NACl with an optimal concentration for activation of about 50 mM. At this ionic strength of 0.05 the chlorides of sodium, potassium, ammonium, rubidium, calcium and magnesium all produced a similar degree of stimulation, while the nitrates of potassium and sodium were somewhat less effective. A similar result was obtained whether the plants were grown in the presence or absence of NACl. Furthermore, malate dehydrogenase activity in preparations from the glycophyte Pisum sativum behaved in a similar manner. The enzyme activity from both Suaeda and Pisum was separable into two fractions (I and II) by gel filtration on Sephadex G200. The MW of fraction II from Suaeda was estimated to be 165000 and that from Pisum approximately 282000: fraction I from both species eluted at the void volume of Sephadex G200. Storage of lyophilised supernatant resulted in the loss of enzyme activity from fraction I and a decrease in the overall stimulation by NaCl. Treatment of the lyophilised enzyme with NACl at a concentration of 100 mM also resulted in the loss of enzyme activity from fraction I.     

A comparison of growth rate of algae as influenced by variation in nitrogen nutrition inChlorella pyrenoidosa andScenedesmus obliquus

Algae were cultivated in nutrient solutions containing nitrates or ammonium salts as the nitrogen source. We measured the culture density in the case of autotrophic and mixotrophic cultivation at different pH values, in the presence of nitrate also at different concentrations and in the absence of molybdenum. The dry weight and amount of nitrogen in the cells under these conditions were determined. According to results obtained we conclude thatChlorella, in contrast toScenedesmus, prefers ammonia as the nitrogen source in the first growth period, later both prefer nitrogen from nitrates. Nitrogen from ammonium salts has a positive effect only during short-time cultivation. If glucose was used as a mixotrophic energy source (or perhaps H-donor), the unsuitability of ammonia as a nitrogen source was more marked. We suppose that the lower growth intensity of older cultures (i.e., after seven days) in ammonia medium is the result of the use of endogenous energy sources. Both algae have adaptive systems making it possible to use nitrate nitrogen even in the absence of molybdate.     

Adverse Effect of Ammonium Salts on the Antibacterial Activity of Paraformaldehyde Solutions

The antibacterial activity of aqueous solutions of paraformaldehyde in concentrations from 0.1 to 0.4% (w/v) is bacteriostatic rather than bactericidal in the presence or absence of ammonium chloride. The presence of ammonium chloride significantly lengthened the time of exposure to paraformaldehyde necessary for inhibition of growth of the test organism (Staphylococcus aureus FDA 209) when unbuffered solutions were used. Elevation of the pH of the reacting mixture of paraformaldehyde and ammonium chloride by partial buffering lengthened the time of exposure necessary for inhibition of growth of the test organism. Decrease of antibacterial activity was concomitant with the disappearance of paraformaldehyde from the reacting mixture. The reaction of paraformaldehyde with ammonium chloride was rapid at room temperature (25 C) and at pH levels near neutrality. The fate of the reacting paraformaldehyde, including the possibility of the formation of hexamethylenetetramine or methylenimine, is discussed with particular reference to loss of antibacterial activity.     

Role of glutamate dehydrogenase and phosphoenolpyruvate carboxylase activity in ammonium nutrition tolerance in roots

Spinach (Spinacea oleracea L. “Correnta F1”) and pea (Pisum sativum L. “Macrocarpon”) plants were grown in a hydroponic culture with nitrate (5 mM), or ammonium (5 mM) as the nitrogen source. Dry matter accumulation declined dramatically in spinach plants fed with ammonium, whereas there was no change in pea plants when compared with nitrate-fed plants. Data obtained from δ¹⁵N, the organic nitrogen content, N-assimilation enzyme activity, glutamine synthetase (L-glutamate:ammonia-ligase; EC 6.3.1.2), glutamate dehydrogenase (L-glutamate:NAD⁺-oxidoreductase; EC 1.4.1.2) and enzymes from the tricarboxylic acid cycle suggest that ammonium incorporation into organic nitrogen is localized in the roots in pea plants and in the shoots in spinach plants. Distribution of incorporated ammonium (in shoots and roots) may determine ammonium tolerance. Our results show that unlike in spinach plants, in pea plants, an ammonium-tolerant species, GDH enzyme plays an important role in ammonium detoxification by its incorporation into amino acids. Furthermore, phosphoenolpyruvate carboxylase (phosphate:oxaloacetate-carboxy-lyase; EC 4.1.1.31) and pyruvate kinase (ATP:pyruvate-2-O-phosphotransferase; EC 2.7.1.40) activities reflect a major flow of carbon for ammonium assimilation through oxalacetate in pea plants and through pyruvate in spinach plants. The differences in the sensitivity to ammonium between the species are discussed in terms of differences in the site of ammonium assimilation as well as in the nitrogen assimilation ways.     

Plant-Induced Hatching of Eggs of the Soybean Cyst Nematode Heterodera glycines

Root diffusate from soybean plants caused greater hatching of Heterodera glycines eggs during vegetative growth of the host, but the activity declined with plant senescence. Chelation of the root diffusate with ethylenediamine tetraacetic acid (EDTA) significantly increased hatching activity for H. glycines eggs. Diffusate from leafless plants caused little hatching, whereas treatment of intact plants with the growth regulators gibberellin and kinetin had no effect on the hatching activity of root diffusate. Treating H. glycines eggs with zinc chloride and root diffusate reduced egg hatching from zinc chloride alone. Levels of zinc in the root diffusate were insufficient to induce egg hatch, based on analysis by atomic absorption spectrophotometry. The enzymatic activity of leucine aminopeptidase in H. glycines eggs was not altered by treatment with chelated or nonchelated root diffusate.     

Replacement of nitrate by ammonium as the nitrogen source increases the salt sensitivity of pea plants. II. Inter- and intracellular solute compartmentation in leaflets

Pea plants (Pisum sativum L. cv. ‘Kleine Rheinländerin’) grown on ammonium or nitrate as the sole nitrogen source were treated with 50 mol m⁻³ NaCl. Four days after salt addition, ammonium-grown plants developed the first visible damage symptoms (wilting of leaflets, starting from the margins). Salt-treated, nitrate-grown plants were not affected during the experimental period. In order to obtain a better understanding of this differential salt sensitivity, we investigated the inter- and intracellular ion compartmentation of leaflets under both nutritional conditions by analysing ion concentrations in the apoplastic space, in chloroplasts and in protoplasts. When the leaves of nitrate- and ammonium-grown plants had attained similar sodium and chloride contents (after different times of exposure to salinity), the latter had a considerably lower chloroplastic chloride (and also sulphate) concentration. The results suggest that the intracellular compartmentation capacity of ammonium-grown plants is considerably lower than that of nitrate-grown plants. Ion toxicity appeared to initiate breakdown of metabolism in parts of the mesophyll tissue of ammonium-grown plants, causing an abrupt release of solutes into the apoplast, which coincided with the appearance of visible damage. Although the ammonium concentrations in leaves increased dramatically in the later phases of damage development, they were too low to cause the collapse of electrochemical gradients at the time at which damage became visible. Thus, the reason for a lower compartmentation capacity under ammonium nutrition remains as yet unclear.     

Catabolite inactivation of phosphoenolpyruvate carboxykinase in spheroplasts from Saccharomyces Cerevisiae

Catabolite inactivation of phosphoenolpyruvate carboxykinase was studied in yeast spheroplasts using 0.9 M mannitol or 0.6 M potassium chloride as the osmotic support. In the presence of potassium chloride the rate of catabolite inactivation was nearly the same as that occurring in intact yeast cells under different conditions of incubation. However, in the presence of mannitol, catabolite inactivation in spheroplasts was prevented. The mannitol inhibition of catabolite inactivation was released by addition of ammonium or phosphate ions. At a concentration of 0.3 M ammonium or 0.06 M phosphate ions, the maximum rate of catabolite inactivation in spheroplasts suspended in mannitol was achieved and was comparable with that observed in spheroplasts incubated in 0.6 M potassium chloride as the osmotic stabilizer. Sodium sulfate (0.04 and 0.4 M) or potassium chloride (0.06 and 0.6 M) did not release the mannitol inhibition of catabolite inactivation in spheroplasts. In intact yeast cells, 0.9 M mannitol, 0.08 M ammonium or 0.1 M phosphate ions did not influence the rate of catabolite inactivation. The nature of the effects of mannitol, ammonium and phosphate ions on catabolite inactivation in yeast spheroplasts is disscussed.     

Effect of selected mercurials on glyceraldehyde-3-phosphate-dehydrogenase in Pteronarcys californica

Glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) was extracted from Pteronarcys californica (Plecoptera) by ammonium sulphate fractionation. The enzyme was treated with three forms of mercury (mercuric chloride, methylmercuric chloride, and phenylmercuric chloride) and their in vitro and in vivo effects on GAPDH were studied. It was found that the orders of toxicity were reversed in the in vivo-in vitro experiments. Electrophoresis was conducted on both treated and untreated enzyme, and showed no differences in mobilities between the two. The enzyme was not found to be inhibited by 0·1 mM iodoacetic acid and required cysteine for activity.     

fus-1, a pH Shift Mutant of Semliki Forest Virus, Acts by Altering Spike Subunit Interactions via a Mutation in the E2 Subunit

Semliki Forest virus (SFV), an enveloped alphavirus, is a well-characterized paradigm for viruses that infect cells via endocytic uptake and low-pH-triggered fusion. The SFV spike protein is composed of a dimer of E1 and E2 transmembrane subunits, which dissociate upon exposure to low pH, liberating E2 and the fusogenic E1 subunit to undergo independent conformational changes. SFV fusion and infection are blocked by agents such as ammonium chloride, which act by raising the pH in the endosome and inhibiting the low-pH-induced conformational changes in the SFV spike protein. We have previously isolated an SFV mutant, fus-1, that requires more acidic pH to trigger its fusion activity and is therefore more sensitive to inhibition by ammonium chloride. The acid shift in the fusion activity of fus-1 was here shown to be due to a more acidic pH threshold for the initial dissociation of the fus-1 spike dimer, thereby resulting in a more acidic pH requirement for the subsequent conformational changes in both fus-1 E1 and fus-1 E2. Sequence analysis demonstrated that the fus-1 phenotype was due to a mutation in the E2 spike subunit, threonine 12 to isoleucine. fus-1 revertants that have regained the parental fusion phenotype and ammonium chloride sensitivity were shown to have also regained E2 threonine 12. Our results identify a region of the SFV E2 spike protein subunit that regulates the pH dependence of E1-catalyzed fusion by controlling the dissociation of the E1/E2 dimer.     

Foliar application of nitrate or ammonium as sole nitrogen supply in Ricinus communis. II. The flows of cations, chloride and abscisic acid

Following a precultivation with pedospheric nitrogen nutrition, Ricinus plants were supplied with nitrogen solely by spraying nitrate or ammonium solution onto the leaves during the experimental period. The chemical composition of tissues, xylem and phloem exudates was determined and on the basis of the previously determined nitrogen flows (Peuke et al., New Phytologist (1998), 138 , 657–687) the flows of potassium, sodium, magnesium, calcium, chloride and ABA were modelled. These data, which permit quantification of net-uptake, transport in xylem and phloem, and utilization in shoot and root, were compared with results obtained in plants with pedospherically-supplied nitrate or ammonium and data in the literature. Although the overall effects on the chemical composition of supplying ammonium to the leaves were not as pronounced as in pedospherically supplied plants, there were some typical responses of plants fed with ammonium (ammonium syndrome). In particular, in ammonium-sprayed plants uptake and transport of magnesium decreased and chloride uptake was increased compared with nitrate-sprayed plants. Furthermore, acropetal ABA transport in the xylem in ammonium-sprayed Ricinus was threefold higher than in nitrate-sprayed plants. Additionally, concentrations of anions were more or less increased in tissues, particularly in the roots, and transport fluids. The overall signal from ammonium-sprayed leaves without a direct effect of ammonium ions on uptake and transport systems in the root is discussed.     

Effect of salts and Triton X-100 on ultrafiltration purification and properties of extracellular glucose oxidase from <Emphasis Type="Italic">Penicillium adametzii</Emphasis> LF F-2044.1

We compared the effectiveness of glucose oxidase isolation from the culture fluid of Penicillium adametzii LF F-2044.1 in the presence of ammonium sulfate, ammonium chloride, and Triton X-100. Ammonium chloride inhibited glucose oxidase in the culture fluid. This compound increased K _M (by 1.2–1.3 times), but decreased V _max for D-glucose oxidation (by 1.7–1.8 times). Ammonium sulfate had little effect on kinetic parameters. Combined treatment with salts and Triton X-100 was followed by a significant increase in the effectiveness of ultrafiltration purification of the culture fluid. The samples of glucose oxidase were electrophoretically characterized. The dependence of kinetic parameters on glucose oxidase concentration during oxidation of D-glucose was evaluated. The catalytic constant and k _cat/K _M ratio for glucose oxidase samples from the culture fluid isolated in the presence of additives significantly surpassed those for enzyme samples, which were obtained by ultrafiltration of the culture fluid with no additives and chromatography on aluminum oxide. The activity of glucose oxidase isolated from the culture fluid in the presence of ammonium chloride was lower compared to that of the enzyme obtained in the presence of ammonium sulfate. This agent is preferable for ultrafiltration of the culture fluid.     

Nitrogen assimilation and cysteine biosynthesis in barley: Evidence for root sulphur assimilation upon recovery from N deprivation

The mixed effects of nitrogen nutrition and sulphate assimilation were investigated in barley plants (Hordeum vulgare var. Alfeo) that were subjected to long-term sulphur and/or nitrogen starvation, by measuring the O-acetylserine(thio)lyase (OASTL-EC 4.2.99.8) activity, changes in -SH compounds and amino acid levels.The growth of barley plants cultured in the hydroponic vessels was severely affected by altered nutrient levels. The barley plants grown in medium deprived of nitrogen and/or sulphur sources for 21 days showed increase in both root length and weight. In contrast, the shoot growth was reduced in nitrogen-starved plants and was unaffected by sulphur deprivation. Sulphur starvation affected the level of proteins in barley plants more than nitrogen deprivation. The decline in the protein levels observed under sulphur-deficient conditions was coupled with the accumulation of glutamine, asparagine and serine, mainly in the roots; additionally, a nitrogen deficiency in the roots promoted a decrease in both glutathione and cysteine levels.The simultaneous deprivation of nitrogen and sulphur in plants leads to an alteration in their metabolism; high levels of glutathione (GSH) in the shoots could signify the induction of a mechanism intended for coping with stressful conditions.Sulphate deprivation enhanced OASTL activity, mainly in the roots; on the other hand, OASTL increases observed under S deprivation were clearly dependent on the nitrogen availability in the culture medium. In fact, the nitrate supply to the N and S starved plants that showed OASTL activity very low, rapidly recovered the OASTL activities to the levels typical of control plants. Nevertheless, the ammonium supply had negligible effects on the OASTL activity only observed after three days in the roots.Our results support the hypothesis that in barley plants, a portion of S assimilation (up to cysteine biosynthesis) occurs in the roots, and a reciprocal influence of nitrogen assimilation on cysteine synthesis occurs.     

Glutamine synthetase regulation by energy charge in sunflower roots

Energy charge [(ATP) + ½ (ADP)]/[(ATP) + (ADP) + (AMP)] and glutamine synthetase activity (transferase reaction) of roots increase in a near congruent manner when decotyledonized sunflower plants (Helianthus annuus L. var. Mammoth Russian) are grown in nitrate for 9 days. Replacement of nitrate with ammonium for the final 2 days leads to a higher energy charge and increased enzyme activity. Similar correlations occur when nitrate plants are placed on a zero nitrogen regimen and when they are subjected to continuous darkness. A rank order correlation of 0.72 is obtained for all data. Control concepts such as adenylylation-deadenylylation and ammonium inhibition of enzyme synthesis are not supported by the data. Energy charge-enzyme activity plots support the view that glutamine synthetase of sunflower roots is subject to control by end products of glutamine metabolism. Alanine appears to exert a modulating effect on the regulation of glutamine synthetase by energy charge.     

Influence of ammonium and nitrate nitrogen on nitrogenase activity of pea plants as affected by light intensity and sugar addition

Summary Addition of ammonium chloride or potassium nitrate to nodulated pea plants resulted in a decrease in acetylene-reducing activity. Both nodule growth and specific activity of the nodules were diminished. Acetylene-reducing activity of isolated bacteroids, treated with EDTA-toluene and supplied with ATP and dithionite, had not decreased after a 3-day treatment of the plants with NH₄Cl or KNO₃. The effect of combined nitrogen could be counteracted by raising the light intensity or by the addition of sucrose to the growth medium. The latter treatment reduced the nitrogen uptake by the plants. It is concluded that combined nitrogen affects symbiotic nitrogen fixation via the carbohydrate supply to the bacteroids.     

Ion Homeostasis in Chloroplasts under Salinity and Mineral Deficiency : I. Solute Concentrations in Leaves and Chloroplasts from Spinach Plants under NaCl or NaNO(3) Salinity

Spinach (Spinacia oleracea var “Yates”) plants in hydroponic culture were exposed to stepwise increased concentrations of NaCl or NaNO₃ up to a final concentration of 300 millimoles per liter, at constant Ca²⁺-concentration. Leaf cell sap and extracts from aqueously isolated spinach chloroplasts were analyzed for mineral cations, anions, amino acids, sugars, and quarternary ammonium compounds. Total osmolality of leaf sap and photosynthetic capacity of leaves were also measured. For comparison, leaf sap from salt-treated pea plants was also analyzed. Spinach plants under NaCl or NaNO₃ salinity took up large amounts of sodium (up to 400 millimoles per liter); nitrate as the accompanying anion was taken up less (up to 90 millimoles per liter) than chloride (up to 450 millimoles per liter). Under chloride salinity, nitrate content in leaves decreased drastically, but total amino acid concentrations remained constant. This response was much more pronounced (and occurred at lower salt concentrations) in leaves from the glycophyte (pea, Pisum sativum var “Kleine Rheinländerin”) than from moderately salt-tolerant spinach. In spinach, sodium chloride or nitrate taken up into leaves was largely sequestered in the vacuoles; both salts induced synthesis of quarternary ammonium compounds, which were accumulated mainly in chloroplasts (and cytosol). This prevented impairment of metabolism, as indicated by an unchanged photosynthetic capacity of leaves.