The effects of amino acids and ammonium on the growth of plant cells in suspension culture

A suspension culture of soybean (Glycine max L.) was grown on a defined medium in which the nitrogen sources were nitrate (25 mM) and ammonium (2 mM). The cells did not grow on nitrate unless the medium was supplemented with ammonium or glutamine. The l- and d-isomers of 12 amino acids tested singly could not replace ammonium. Most amino acids (4 mM) inhibited growth when the cells were cultured on nitrate and ammonium. Cells from five other plants (Reseda luteoli L.; Triticum monococcum L.; flax, Linum usitatissimum L.; horseradish, Amoracia lapathifolia Gilib; Haplopappus gracilis L.) grew on the defined medium with nitrate (25 mM) as the sole nitrogen source. Higher cell yields were obtained when ammonium (2 mM) or glutamine also was present. Supplementing the defined medium with high concentrations of ammonium (20 mM) inhibited growth of soybean, Haplopappus, and wheat cells. Addition of citrate (5 mM) relieved the inhibitory effects of ammonium in soybean and wheat cells but not in the Haplopappus cells.     

Carrier-mediated Uptake of Arginine and Urea by Chlamydomonas reinhardtii

Chlamydomonas reinhardtii possesses a high affinity, highly specific carrier involved in uptake of exogenous arginine. Carrier-mediated uptake of other amino acids cannot be detected, even in cultures maintained on amino acids as a nitrogen source or starved for nitrogen. This fact may contribute to the difficulty of isolating strains auxotrophic for amino acids other than arginine; conventional selection media may not supply adequate quantities of amino acids to permit growth of auxotrophs. A urea carrier is also present in C. reinhardtii but is readily distinguished from the arginine carrier on the basis of kinetic properties and sensitivity to a range of structural analogs. Ammonia appears to play a major role in regulating (depressing) activity of the arginine uptake system. Activity of the urea uptake system is elevated in nitrogen-starved cultures and elevated even further in the presence of urea or arginine. Extensive, independent fluctuations in the two uptake systems observed in semisynchronous cultures suggest that both are subject to modulation by a complex set of interacting endogenous and exogenous factors.     

Inhibition of cultured cell growth by tungstate and molybdate

The growth of suspension cultured cells of Nicotiana tabacum (tobacco) was inhibited completely by 100 M tungstate. Even though molybdate reversed the tungstate inactivation of nitrate reductase activity, the growth inhibition was not reversed. The growth inhibition of N. tabacum, Daucus carota, Glycine max and Solanum tuberosum suspension cultured cells by tungstate was similar in media with or without amino acids as a source of reduced nitrogen. Only in the case of G. max was a slight reversal caused by the amino acids. Tungstate was slightly less inhibitory to the growth of a nitrate reductase-lacking mutant N. tabacum line (nia-63) than to the line with nitrate reductase. These results indicate that tungstate must inhibit the cell growth of the four species used, predominantly, in some way other than by inhibiting nitrate reductase activity. Similar studies with molybdate, a sulfate analog which apparently competes with sulfate at the ATP sulfury-lase enzyme, showed that 1 mM concentrations were completely inhibitory to cell growth. The addition of sulfate or cysteine, as a source of reduced sulfur, and amino acids, as a source of reduced nitrogen, in most cases did not reverse the molybdate inhibition appreciably. Some reversal was seen only by sulfate with D. carota cells and by cysteine plus amino acids with D. carota and G. max. These results indicate that selection for tungstate or molybdate resistance will in general not select for higher levels or other alterations in the activity of nitrate reductase or ATP sulfurylase, respectively, since these ions do not inhibit growth by primarily affecting these enzymatic steps in cultured cells of the four species studied.     

Effect of growth conditions on accumulation of internal nitrate,ammonium, amino acids,and protein in three marine diatoms

The capacity of marine phytoplankton to change their cellular content of nitrate, ammonium, amino acids, and protein in response to different growth conditions was systematically investigated. Cellular concentrations of these compounds were measured in N-starved, N-deficient, and N-sufficient Skeletonema costatum (Grev.) Cleve and in N-deficient Chaetoceros debilis Cleve and Thalassiosira gravida Cleve, both before and after the addition of a pulse of nitrogen.N-sufficient Skeletonema costatum contains high concentrations of protein, large persistent pools of amino acids, and, if it is growing on nitrate, sizeable amounts of nitrate. As it becomes N-starved, the total cellular nitrogen decreases, the internal nitrate and amino acids become entirely depleted, and the protein content is drastically reduced. After nitrogen additions to N-deficient and N-starved cultures, transient pools of unassimilated nitrogen form which can account for a large fraction of newly taken up nitrogen. The size and kind of pool which accumulates is determined by the preconditioning of the cells, the nitrogen compound which is added, and the species identity. The pools which form in S. costatum indicate that nitrate reduction is the slowest step in nitrogen assimilation, the synthesis of protein from amino acids is the next slowest, and the incorporation of ammonium into amino acid is the fastest. However, the rate limiting steps may vary between diatom species.For the first time, measurements of the variation in cellular nitrogen compounds over a wide range of environmental conditions reveal the ability of some phytoplankton to buffer the effects of a changing, and sometimes growth-limiting, nitrogen supply. They accomplish this by utilizing stored internal nitrogen for growth when the external supply is low and by quickly storing unassimilated nitrogen when the external supply is suddenly increased beyond their ability to immediately assimilate it. The accumulation of large pools of unassimilated nitrogen compounds can explain the often observed difference between nitrogen uptake rates and growth rates.     

Differential regulation of nitrate reductase induction in roots and shoots of cotton plants

The induction of nitrate reductase activity in root tips of cotton (Gossypium hirsutum L.) was regulated by several amino acids and by ammonium. Glycine, glutamine, and asparagine strongly inhibited induction of activity by nitrate and also decreased growth of sterile-cultured roots on a nitrate medium. Methionine, serine, and alanine weakly inhibited induction, and 11 other amino acids had little or no effect. Ammonium also decreased induction in root tips, but was most effective only at pH 7 or higher. The optimum conditions for ammonium regulation of induction were identical to those for growth of sterile-cultured roots on ammonium as the sole nitrogen source. Aspartate and glutamate strongly stimulated induction, but several lines of evidence indicated that the mechanism of this response was different from that elicited by the other amino acids. The effects of amino acids on induction appeared to be independent of nitrate uptake.     

Nitrogen source and mineral optimization enhance d-xylose conversion to ethanol by the yeast Pichia stipitis NRRL Y-7124

Nutrition-based strategies to optimize xylose to ethanol conversion by Pichia stipitis were identified in growing and stationary-phase cultures provided with a defined medium varied in nitrogen, vitamin, purine/pyrimidine, and mineral content via full or partial factorial designs. It is surprising to note that stationary-phase cultures were unable to ferment xylose (or glucose) to ethanol without the addition of a nitrogen source, such as amino acids. Ethanol accumulation increased with arginine, alanine, aspartic acid, glutamic acid, glycine, histidine, leucine, and tyrosine, but declined with isoleucine. Ethanol production from 150 g/l xylose was maximized (61±9 g/l) by providing C:N in the vicinity of ∼57–126:1 and optimizing the combination of urea and amino acids to supply 40–80 % nitrogen from urea and 60–20 % from amino acids (casamino acids supplemented with tryptophan and cysteine). When either urea or amino acids were used as sole nitrogen source, ethanol accumulation dropped to 11 or 24 g/l, respectively, from the maximum of 46 g/l for the optimal nitrogen combination. The interaction of minerals with amino acids and/or urea was key to optimizing ethanol production by cells in both growing and stationary-phase cultures. In nongrowing cultures supplied with nitrogen as amino acids, ethanol concentration increased from 24 to 54 g/l with the addition of an optimized mineral supplement of Fe, Mn, Mg, Ca, Zn, and others.The mention of trade names or commercial products in this article is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture.     

Submerged culture conidial germination and conidiation of the bioherbicideColletotrichum truncatum are influenced by the amino acid composition of the medium

Summary Submerged culture experiments were conducted to determine the optimal nitrogen source for rapidly producing conidia of the bioherbicide,Colletotrichum truncatum. Germination ofC. truncatum conidial inocula in submerged culture occurred most rapidly (>95% in 6 h) in media provided with a complete complement of amino acids. When (NH₄)₂SO₄, urea, or individual amino acids were provided as the sole nitrogen source, conidial germination was less than 20% after 6 h incubation. Conidia production was delayed inC. truncatum cultures grown in media with urea or individual amino acids as nitrogen sources compared to cultures supplied with Casamino acids or complete synthetic amino acid nitrogen sources. The use of methionine, lysine, tryptophan, isoleucine, leucine or cysteine as a sole nitrogen source severely inhibitedC. truncatum conidia production. Media with synthetic amino acid mixtures less these inhibitory amino acids produced significantly higher conidia yields compared to media with amino acid mixtures containing these amino acids. When various amounts of each individual inhibitory amino acid were added to media which contained amino acid mixtures, cysteine and methionine were shown to be most effective in reducing conidiation. An optimal nitrogen source forC. truncatum conidiation in submerged culture should contain a complete mixture of amino acids with low levels of cysteine, methionine, leucine, isoleucine, lysine and tryptophan for rapid conidiation and optimal conidia yield.The mention of firm names or trade products does not imply that they are endorsed or recommended by the US Department of Agriculture over other firms or similar products not mentioned.     

Characterization of urease from the phototrophic bacteriumRhodobacter capsulatus E1F1

Rhodobacter capsulatus E1F1 showed high cytosolic urease activity when growing on urea, purines, and purine metabolites as nitrogen source. Molecular mass ofR. capsulatus enzyme is similar to that of other bacteria and greatly differs from that of jack bean. Kinetic parameters of partially purifiedR. capsulatus enzyme resemble those described in other bacterial ureases. The activity was inhibited by metal-chelating agents and by mercurials. Urease fromR. capsulatus E1F1 was negligible in nitrogen-starved cells or in cells cultured with nitrate, ammonium, or amino acids. Moreover, ammonium inhibited both the urea uptake and the urease activity expression inR. capsulatus cells.     

CHEMOSENSORY RESPONSES OF THE SYMBIOTIC DINOFLAGELLATE SYMBIODINIUM MICROADRIATICUM (DINOPHYCEAE)1

Motile Symbiodinium microadriaticum (Freudenthal 1962) were attracted to a variety of nitrogen-containing compounds, including ammonium, nitrate, urea and some amino acids. No chemosensory response to phosphate, sulphate, vitamins, trace metals or sugars was evident. Motile algae responded to concentrations of ammonium, nitrate, and urea at least as low as 10^?6 M. High concentrations (≥ 10^?2 M) of ammonium appeared to inhibit attraction of motile algae. Calculations using ammonium release rates from various aposymbiotic hosts suggest that motile S. microadriaticum can respond to released ammonium ca. 1 cm from the source. Cultured algae were not attracted to combined nitrogen cues for at least 2 days after inoculation into seawater with dissolved combined low nitrogen. Algae freshly isolated from starved animals were normally motile the day following isolation and attracted to ammonium and nitrate when maintained in seawater containing < 1 μM ammonium and nitrate. The algae lost their ability to orient to nitrogen attractants the day after incubation into culture medium containing high levels of ammonium and nitrate. These results suggest that chemosensory behavior is suppressed when nutrients are present in the ambient medium or are stored by the alga. There were few differences in chemosensory abilities in different strains of S. microadriaticum to the attractants assayed, suggesting that selection for a particular strain by a host species may not be due to differential chemosensory ability or cues. However, the absence of chemical attraction of motile S. microadriaticum to infected hosts may act to preserve strain selection occurring at other steps in the infection process of aposymbiotic hosts.     

Effects of amino acids, nitrate, and ammonium on the growth and taxol production in cell cultures of Taxus yunnanensis

The effects of concentration of amino acids, nitrate, and ammonium on the growth and taxol production in cultures of cell line TY-21 of Taxus yunnanensis were investigated. Addition of 20 different amino acids each at 15–20 mg l^–1 to B5 medium significantly improved callus growth but inhibited taxol formation in the cultures. The optimum nitrate concentration was 20–30 mM for both growth and taxol production. Ammonium greatly suppressed growth but strongly promoted taxol formation in the cells when it was the sole inorganic nitrogen in the medium. Culturing the suspension cells in nitrate-containing medium for 15 days and then in a medium in which ammonium was the sole inorganic nitrogen for 7 days increased taxol yield by 104%, reaching up to 28.1 mg l^–1.     

Effect of glyphosate on carrot and tobacco cells

The growth of suspension-cultured carrot (Daucus carota L.) and tobacco (Nicotiana tabacum L. cv. Xanthi) cells was inhibited by glyphosate (N-[phosphonomethyl]glycine). This inhibition was reversed by adding combinations of phenylalanine, tyrosine, and tryptophan or casein hydrolysate. Casein hydrolysate and phenylalanine + tyrosine + tryptophan were the most effective treatments. Reversal of glyphosate-induced inhibition occurred only if the aromatic amino acids were added during the first 8 days of glyphosate incubation. Glyphosate uptake was not reduced when the aromatic amino acids or casein hydrolysate were added.     

Amino Acid interactions in the regulation of nitrate reductase induction in cotton root tips

Glycine, asparagine, and glutamine inhibited the induction by nitrate of nitrate reductase activity in root tips of cotton (Gossypium hirsutum L.). This inhibition was partially or entirely prevented when the inhibitor was applied in combination with any of several other amino acids. Studies of ¹⁴C-labeled amino acid uptake showed that, in most cases, the apparent antagonism resulted simply from competition for uptake. However, certain antagonists did not curtail uptake. The most effective of these were leucine (against all three inhibitors), and isoleucine and valine (against asparagine or glutamine, but not glycine). These results show that interactions among amino acids in the regulation of nitrate reductase induction result from at least two mechanisms, one acting on uptake of inhibitory amino acids, and the other involving true antagonism.     

Distribution of Nitrogen during Growth of Sunflower (Helianthus annuus L.)

The accumulation, distribution and redistribution of dry matterand nitrogen is described for Helianthus annuus L. cv. Hysun21 grown on 6 mM urea in glasshouse culture. Seed dry matterand nitrogen were transferred to seedlings with net efficienciesof 40 and 86 per cent respectively. At flowering, the stem hadmost of the plant's dry matter and the leaves most of its nitrogen.About 35 per cent of the plant's nitrogen accumulated afterthree-row anthesis. The amount of protein in vegetative parts,especially leaves, declined after flowering. Concentrationsof free amino compounds also decreased during growth. Matureseeds had 38 per cent of the total plant dry weight and 68 percent of the total nitrogen. Seeds acquired 33 per cent of theirdry matter and nitrogen from redistribution from above-groundplant parts. The stem was most important for storage of carbohydrate,leaves the most important for nitrogen. Over 50 per cent ofthe nitrogen in the stem and leaves was redistributed. Plantsthat received 6 mM nitrate accumulated more dry matter thanurea-grown plants. Seeds from nitrate-grown plants were heavier(58 mg) than those of urea-grown plants (46 mg), and their percentageoil was greater (50 and 41 respectively). The amount of nitrogenper seed was the same. Little or no urea was detected in xylem sap of plants suppliedwith 5 mM urea, but it was detected in sap of plants which received25 mM. Concentrations of urea and amino compounds in the sapdecreased up the stem. Plants supplied with nitrate had mostof the nitrogen in xylem sap as NO₂^–, suggesting littlenitrate reduction in roots. Plants grown on 6 mM nitrate andchanged to high levels of urea-nitrogen for 14 days still hadhigh levels of nitrate; little nitrate remained in plants receivinglow levels of urea. When urea is applied in irrigation waterto field-grown sunflower, the nitrogen is subsequently takenup as nitrate due to rapid nitrogen transformations in the soil. Helianthus annuus L., sunflower, urea, nitrate, nitrogen transport, xylem sap, nitrogen accumulation nitrogen distribution     

Nitrate Assimilation in Higher Plants with Special Reference to the Cocklebur (Xanthium pennsylvanicum Wallr.)

A soluble NADH-dependent nitrate reductase is described forthe shoot system of Xanthium. Young leaves and immature stemtissues contain high levels of the enzyme. They are relativelyrich in free amino acids and amides but store little free nitrate.The specific activity of the enzyme is lower in fully expandedleaves, although these leaves exhibit higher rates of fixationof carbon in photosynthesis than do younger leaves. Neithernitrate nor free amino acids accumulate in the mesophyll ofthe leaf. Older parts of the stem axis accumulate large amountsof soluble nitrogen, almost entirely as free nitrate. Reservesof nitrate in the shoot and root are rapidly depleted if nitrateis removed from the external medium. Nitrate reductase is apparently absent from roots of Xanthium.This finding is supported by analyses of bleeding sap from nitrate-fedplants which show that 95 per cent of the nitrogen exportedfrom roots is present as free nitrate. However, roots are capableof synthesizing and exporting large amounts of amino nitrogenif supplied with reduced nitrogen such as urea or ammonium. A scheme is presented summarizing the main features of the metabolismof nitrate in Xanthium and this is compared with the situationin nitrate-fed plants of the field pea (Pisum arvense L.), aspecies previously shown to be capable of reducing nitrate inits root system.     

Nitrate reductase activity and growth in Paul's Scarlet rose suspension cultures in relation to nitrogen source and molybdenum

Growth and nitrate reductase activity were measured in Paul's Scarlet rose cell suspensions, cultured in media purified from molybdenum and containing nitrate or urea as sole nitrogen source with or without added Mo. Urea could replace nitrate to yield 80% of the fresh weight in nitrate medium. Nitrate reductase activities were compared by in vivo and in vitro assays. The latter varied due to inactivation during extraction. Compared with activities in cells in complete NO₃ ^- medium, activity in NO₃ ^--Mo cells was reduced to 30% and, in urea-grown cells, to trace amounts. Increases in nitrate reductase activity were found when NO₃ ^- alone was added to NO₃ ^- or urea+Mo cultures. In NO₃ ^--Mo cultures, Mo alone or with NO₃ ^- caused a similar increase in activity, whereas urea-Mo cultures required both NO₃ ^- and Mo for enzyme induction.Abbreviations FAD flavin adenine dinucleotide - Mo molybdenum - NADH reduced nicotinamide adenine dinucleotide - NO₃ ^-+Mo standard MX₁ culture medium - NO₃ ^--Mo MX₁ medium purified of Mo and used for continuous subculture with nitrate - NR nitrate reductase - PSR Paul's Scarlet rose - PVP polyvinylpyrrolidone - U urea - U+Mo MX₁ medium containing urea instead of nitrate - U-Mo MX₁ medium containing urea instead of nitrate and also purified of Mo     

Growth response of four freshwater algal species to dissolved organic nitrogen of different concentration and complexity

相似文献   

Blue-light-control of the uptake of amino acids and of ammonia in Chlorella mutants

In non-photosynthetic, yellow or colourless mutant cells of Chlorella kessleri , grown with nitrate as sole nitrogen source, blue light inhibited the uptake of the amino acids glycine, proline and arginine and of ammonia in growing cells, while it enhanced the uptake of these amino acids in resting cells. On the other hand, in cells grown with ammonia as the only nitrogen source without nitrate reductase activity, blue light did not influence the uptake of amino acids and of ammonia in growing cells, while it enhanced the uptake of amino acids in resting cells. Addition of methionine sulphoximine, a potent inhibitor of glutamine synthetase, to growing cells, resulted in intracellular ammonia-accumulation and inhibition of uptake of glycine and of ammonia. For the colourless mutant, blue light was shown to activate purified nitrate reductase. These results indicate that in the mutant cells of Chlorella examined, uptake of ammonia seems to be influenced by nitrate reductase and the uptake of amino acids was influenced by both nitrate reductase and an unknown blue-light-receptor(s). The uptake of urea in mutant cells is not influenced by the irradiation with blue light. Uptake of glycine was also increased after addition of glucose (hexose) in the dark. Because blue light is known to enhance the breakdown of starch, a reaction producing glucose for oxidative degradation in the algae used, the role of glucose (hexose) in the blue light-affected uptake of amino acids is discussed.     

Nitrogen Assimilation in Barley (Hordeum vulgare L. cv. Mazurka) in Response to Nitrate and Ammonium Nutrition

Barley plants (Hordeum vulgare L. cv. Mazurka) were grown inaerated solution cultures with 2 mM or 8 mM inorganic nitrogensupplied as nitrate alone, ammonium alone or 1:1 nitrate+ammonium.Activities of the principal inorganic nitrogen assimilatoryenzymes and nitrogen transport were measured. Activities ofnitrate and nitrite reductases, glutamine synthetase and glutamatesynthase were greater in leaves than in roots but glutamatedehydrogenase was most active in roots. Only nitrate and nitritereductases changed notably (4–10 times) in response tothe different nitrogen treatments. Nitrate reductase appearedto be rate-limiting for nitrate assimilation to glutamate inroots and also in leaves, where its total in vitro activitywas closely related to nitrate flux in the xylem sap and wasslightly in excess of that needed to reduce the transportednitrate. Xylem nitrate concentration was 13 times greater thanthat in the nutrient solution. Ammonium nitrogen was assimilatedalmost completely in the roots and the small amount releasedinto the xylem sap was similar for the nitrate and the ammoniumtreatments. The presence of ammonium in the nutrient decreasedboth export of nitrate to the xylem and its accumulation inleaves and roots. Nitrate was stored in stem bases and was releasedto the xylem and thence to the leaves during nitrogen starvation.In these experiments, ammonium was assimilated principally inthe roots and nitrate in the leaves. Any advantage of this divisionof function may depend partly on total conversion of inorganicnitrogen to amino acids when nitrate and ammonium are givenin optimal concentrations. Hordeum vulgare L., barley, nitrate, ammonium, nitrate reductase, nitrite reductase, glutamine synthetase, glutamate synthase, glutamate dehydrogenase, nitrogen transport