Aspects of inorganic nitrogen assimilation in yeasts

Cultures of Candida utilis utilise glutamate in preference to ammonia and ammonia in preference to nitrate. The nitrate reductase of this organism is induced by nitrate and repressed in cultures grown on glutamate or ammonia. Nitrate-grown cultures of C. utilis, irrespective of the medium nitrate concentration, behave as though nitrogen-limited. In contrast to C. utilis, Saccharomyces cerevisiae utilises ammonia in preference to glutamate. In eight yeasts studied the highest cellular contents of biosynthetic NADP-linked glutamate dehydrogenase were found in batch cultures containing low concentrations of ammonia or in nitrogen-limited chemostat cultures. NAD-linked glutamate dehydrogenase activity was detected in extracts of cells grown in the presence of glutamate but not in those grown in the presence of ammonia.     

Temperature dependence of inorganic nitrogen uptake: reduced affinity for nitrate at suboptimal temperatures in both algae and bacteria.

Nitrate utilization and ammonium utilization were studied by using three algal isolates, six bacterial isolates, and a range of temperatures in chemostat and batch cultures. We quantified affinities for both substrates by determining specific affinities (specific affinity = maximum growth rate/half-saturation constant) based on estimates of kinetic parameters obtained from chemostat experiments. At suboptimal temperatures, the residual concentrations of nitrate in batch cultures and the steady-state concentrations of nitrate in chemostat cultures both increased. The specific affinity for nitrate was strongly dependent on temperature (Q10 approximately 3, where Q10 is the proportional change with a 10 degrees C temperature increase) and consistently decreased at temperatures below the optimum temperature. In contrast, the steady-state concentrations of ammonium remained relatively constant over the same temperature range, and the specific affinity for ammonium exhibited no clear temperature dependence. This is the first time that a consistent effect of low temperature on affinity for nitrate has been identified for psychrophilic, mesophilic, and thermophilic bacteria and algae. The different responses of nitrate uptake and ammonium uptake to temperature imply that there is increasing dependence on ammonium as an inorganic nitrogen source at low temperatures.     

Molecular analysis of inorganic nitrogen assimilation in yeasts

Assimilation of nitrate and various other inorganic nitrogen compounds by different yeasts was investigated. Nitrate, nitrite, hydroxylamine, hydrazine, ammonium sulphate, urea and L-asparagine were tested as sole sources of nitrogen for the growth of Candida albicans, C. pelliculosa, Debaryomyces hansenii, Saccharomyces cerevisiae, C. tropicalis, and C. utilis. Ammonium sulphate and L-asparagine supported the growth of all the yeasts tested except D. hansenii while hydroxylamine and hydrazine failed to support the growth of any. Nitrate and nitrite were assimilated only by C. utilis. Nitrate utilization by C. utilis was also accompanied by the enzymatic activities of NAD(P)H: nitrate oxidoreductase (EC 1.6.6.2) and NAD(P)H: nitrite oxidoreductase (EC 1.6.6.4), but not reduced methyl viologen-or FAD-nitrate oxidoreductases (EC 1.7.99.4). It is demonstrated here that nitrate and nitrite reductase activities are responsible for the ability of C. utilis to assimilate primary nitrogen.     

Nitrogenous nutrition of sea-ice microalgae

Summary There are indications that the final biomass attained by sea-ice microalgae in southeastern Hudson Bay is nitrogen limited. The present study investigates the possibility that the rate at which the final yield is approached is also nitrogen limited. Nutrient data suggest that nitrogen was actively taken up by the microalgae, and periodically replenished by mixing processes related to fortnightly tides. High molar C:N ratios (>15), typical of nitrogen-deficient cells, have been observed at times of low dissolved inorganic nitrogen. Absolute transport rates of nitrogen were estimated from natural changes in dissolved inorganic and particulate organic nitrogen. Cell losses and nitrogen uptake rates were highest during periods of maximum current speed, suggesting that the rate of biological production at the ice-water interface could be limited by the accumulated biomass. These results suggest (1) that the sea-ice microalgae in southeastern Hudson Bay are nitrogen limited in their natural environment, and (2) that nutrient replenishment and perhaps losses of biomass governed by fortnightly tidal mixing periodically enhance the growth of microalgae at the ice-water interface.Contribution to the programs of the Maurice Lamontagne Institute and of the Groupe interuniversitaire de recherches océanographiques du Québec (GIROQ)     

Inorganic carbon uptake by an Antarctic sea-ice diatom,Nitzschia frigida

There have been no studies to date on the mechanisms of inorganic carbon acquisition by Antarctic microalgae. Consequently, we have examined inorganic carbon (DIC) use inNitzschia frigida, a diatom typical of the Antarctic bottom-ice community. The K_0.5(CO₂) of photosynthesis in this organism was estimated to be 1.09 μM at pH 7.5. The internal concentration of DIC was approximately 4050 μM at an external [DIC] of 45 μM. At air-equilibration levels of inorganic carbon this would be sufficient for a ten-fold accumulation ratio of CO₂. Cells ofN. frigida are capable of carbon-dependent photosynthesis at rates that exceed that expected from uncatalysed CO₂ supply to the cell. About 25% of the total carbonic anhydrase activity appears to be associated with the cell surface inN. frigida. These results support the hypothesis thatN. frigida, like many microalgae from temperate waters, has an active carbon-concentrating mechanism, associated with the ability to utilize external HCO ₃ ⁻ for photosynthesis.     

Organic and inorganic nitrogen uptake in lichens

In order to learn more about nitrogen (N) acquisition in lichens, and to see whether different lichens differ in their affinity to various N sources, N uptake was measured in 14 various lichen associations (species). These species represented various morphologies (fruticose or foliose), contrasting microhabitat preferences (epiphytic or terricolous), and had green algal, cyanobacterial or both forms of photobionts. N was supplied under non-limiting conditions as an amino acid mixture, ammonium, or nitrate, using ¹⁵N to quantify uptake. Carbonyl cyanide m-chlorophenylhydrazone (CCCP) was used to separate active and passive uptake. Thallus N, amino acids, soluble polyol concentrations, and the biont-specific markers chlorophyll a and ergosterol were quantified, aiming to test if these metabolites or markers were correlated with N uptake capacity. Ammonium uptake was significantly greater and to a higher extent passive, relative to the other two N sources. Nitrate uptake differed among lichen photobiont groups, cyanobacterial lichens having a lower uptake rate. All lichens had the capacity to assimilate amino acids, in many species at rates equal to nitrate uptake or even higher, suggesting that organic N compounds could potentially have an important role in the N nutrition of these organisms. There were no clear correlations between N uptake rates and any of the measured metabolites or markers. The relative uptake rates of ammonium, nitrate and amino acids were not related to morphology or microhabitat.Abbreviations CCCP Carbonyl cyanide m-chlorophenylhydrazone - Chl Chlorophyll - N Nitrogen     

Seasonal periodicity of physical factors,inorganic nutrients and microalgae in Antarctic fellfields

Over 15 months between January 1990 and March 1991, a range of physical, chemical and biological parameters was monitored regularly in fellfield soils of frost-sorted polygons at four sites on Signy Island (South Orkney Islands, maritime Antarctica). These included inorganic nutrients (orthophosphate, available nitrate and, over a more limited period, ammonia), chlorophyll a (as a proxy measure of microalgal biomass) and a range of potential cryoprotectant compounds. Transects across soil polygons revealed neither intrapolygon gradients in concentrations of inorganic nutrients or chlorophyll a nor significant interpolygon differences, in contrast with previous studies. Nitrate was present in much lower concentrations than phosphate, supporting evidence that it is a limiting nutrient in these fellfield ecosystems. Spring snowmelt, although a potential source of nutrient input, was not associated with increased concentrations of inorganic nutrients in the soil, probably through isolation of the soil from overlaying snow by a surface layer of ice. Soil microalgae at the study sites must survive winter temperatures of at least −9°C, even when protected beneath up to 1 m of snow, and it has been proposed that they accumulate sugars and polyols as cryoprotectants. In support of this, concentrations of erythritol, glycerol, glucose, sucrose and trehalose in the soil (both absolute quantity and after correction for chlorophyll a concentration) increased as winter proceeded, suggesting that changes in sugar concentrations were due to accumulation within individual cells.     

Photoadaptation of sea-ice microalgae in the Barents Sea

Summary Variations in under-ice scalar irradiance, P vs I parameters and the CHLa C^–1 ratio of natural assemblages of sea-ice microalgae from the Barents Sea growing at -1.8°C in May and September 1988 are described, including one diurnal station. CHLa C^–1 ratios of 0.031–0.071 mg mg^–1 indicate shade adaptated assemblages both in May and September. Values for ^B (photosynthetic efficiency) were generally low, e.g. 0.0025–0.0078 mg C (mg CHLa)^–1 h^–1 (mol m^–2 s^–1)^–1, and should be typical for self-shaded algae in mats or aggregates of about 4 mm thickness. Provided no self shading and the typical spectral distribution of light under ice without algae, ^B would, however, be about 2.5 times higher. Photoinhibition of the photosynthetic response was negligible. Maximum carbon uptake P _m ^B was 0.15–0.24 and 0.032–0.088 mg C (mg CHLa)^–1 h^–1 in May and September, respectively. Diurnal variations were small, particularly for P _m ^B . Calculations of the maximum specific gross growth rate yielded an upper limit of 0.20–0.24 and 0.01–0.04 d^–1 for assemblages in May and September, respectively; the latter may have been in a resting stage.Contribution No. 245, Trondhjem Biological Station     

Characteristics of inorganic nitrogen assimilation of plants in fire-prone Mediterranean-type vegetation

A range of approaches was used to investigate how species within a fire-prone Banksia woodland in South West Australia exploited inorganic soil nitrogen sources and how this changes through the development of the fire chronosequence. Nitrate and ammonium were present in soil solution throughout the chronosequence but nitrate predominated in recently burnt sites. Mean shoot nitrate reductase activities were high for all species in recently burnt sites and showed little increase when nitrate was supplied via the transpiration stream. Nitrate reductase of shoots of most species was low at sites not burnt for several years, but following transpirational induction with nitrate, developed activities similar to those at recently burnt sites. The principal amino compounds transported in the xylem were species specific, including asparagine, glutamine and citrulline-dominated species, and changed little in relative composition across the chronosequence. Species most active in leaf nitrate reduction transported the largest amounts of nitrate in their xylem sap and proportional amounts of nitrate in xylem tended to be greatest in recently burnt sites. Most of the species examined appeared to be shoot rather than root nitrate assimilators, but marked differences were recorded in potential of leafy shoots of different species to reduce nitrate. As a general rule, shallow-rooted herbaceous, non-mycorrhizal or VAM-positive species had the highest capacity to reduce nitrate, whereas woody species with ericoid mycorrhizae or combined vesicular arbuscular/ectomycorrhizal associations exhibited little capacity to reduce nitrate in roots or shoots. It seems likely that this latter group utilize ammonium or even organic forms of nitrogen rather than nitrate. Some putative nitrogen-fixing species were active in reducing and transporting nitrate, others were virtually inactive in these respects.     

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.     

Diel changes of uptake of inorganic carbon and nitrogen by phytoplankton, and the relationship between inorganic carbon and nitrogen uptake in Lake Nakanuma, Japan

Diel changes of uptake of inorganic carbon and nitrogen wereexamined in a small freshwater lake, Lake Nakanuma, Japan, bythe ¹³C and ¹⁵N method. Experiments were earned out in spring,summer and autumn in 1984. Carbon and nitrogen uptake in thelight incubation showed maxima around noon at the three seasons.Carbon uptake ceased at night, but ammonium uptake was fairlylarge at night. In the dark incubation carbon uptake did notoccur. Ammonium uptake showed a maximum at dusk in the darkexperiments. Diel changes of nitrate uptake were less clearthan those of ammonium uptake. These results indicate that nitrogenuptake partly depended on the carbon uptake. Then, we triedto explain the diel changes of nitrogen uptake, assuming thatthe nitrogen uptake partly depends on stored carbohydrate. Thediel changes may be elucidated by the sum of three terms: oneis the term of decay of stored carbohydrate, the second is theterm which indicates cumulative increase of stored carbohydrateand the third is the term which directly depends on light.     

Relationships between nitrogen uptake and carbon assimilation in whole plants of tall fescue

Abstract. The present study investigates the relationships between nitrogen uptake, transpiration, and carbon assimilation. Plants growing on nutrient solution were enclosed for 10–16 d in a growth chamber, where temperature, photon flux density, vapour saturation deficit and CO₂ concentration were controlled. One of these factors was modified every 4 to 5 d. Shoot photosynthesis and root and shoot respiration were recorded every half-hour. Nitrogen uptake from the root medium and plant transpiration were measured daily. In most cases, an increase in photon flux density led to increases in transpiration, net daily carbon assimilation, and nitrogen uptake. By modifying transpiration rate without changing photosynthesis (varying vapour saturation deficit), or by modifying transpiration and carbon assimilation in opposite ways (varying CO₂ air concentration), it was shown that nitrogen uptake does not follow transpiration, but is linked to the carbon uptake of the plant. When light was increased from low to intermediate levels, the N uptake/C assimilation ratio remained constant. At higher photon flux density, this ratio declined markedly. It is proposed that in the first case, growth is limited by carbohydrate availability, thus any increase in carbon assimilation leads to a proportional increase in nitrogen uptake, in contrast to the second situation where carbohydrates may accumulate in the plant without further nitrogen requirement.     

Availability and uptake of inorganic nitrogen in a mixed old-growth coniferous forest

Old-growth forest stands of mixed species composition provide the opportunity to study species-specific influences on soil properties. We monitored rates of nitrogen mineralization, nitrification and an index of ammonium and nitrate uptake in a mixed old-growth stand of Douglas-fir (Pseudotsuga menziesii), western hemlock (Tsuga heterophylla) and western redcedar (Thuja plicata) over a two-year period. Litter and mineral soil (0–10-cm depth) were sampled adjacent to ten large trees of each species. After initial characterization of litter and soil, buried bags were incubated in both layers for ca. 2-month intervals. Soil and litter pH was lowest near western hemlocks. Nitrification, nitrate concentrations, and percent uptake as nitrate differed among the tree species; rates were highest near western redcedars. For all species, percent nitrification and nitrate uptake rate were higher in soil than in litter. The results indicate species-specific effects on ammonium and nitrate production and uptake within this forest type.The research described in this article has been funded in part by the US Environmental Protection Agency. This document has been prepared at the EPA Environmental Research Laboratory in Corvallis, Oregon, through Contract No. 68-C8-0006 to ManTech Environmental Technology, Inc. It has been subjected to the Agency's peer and administrative review and approved for publication. Mention of trade names or commercial products does not constitute endorsement or recommendation for use.The research described in this article has been funded in part by the US Environmental Protection Agency. This document has been prepared at the EPA Environmental Research Laboratory in Corvallis, Oregon, through Contract No. 68-C8-0006 to ManTech Environmental Technology, Inc. It has been subjected to the Agency's peer and administrative review and approved for publication. Mention of trade names or commercial products does not constitute endorsement or recommendation for use.     

Photosynthetic responses of Arctic sea-ice microalgae to short-term temperature acclimation

Summary In April-May 1986, sea-ice microalgae (southcastern Hudson Bay, Canadian Arctic) were acclimated to temperatures ranging from-1.5° to 10°C for short periods (3 h), after which photosynthesis and carboxylating enzyme activities were measured. P _max ^b increased after acclimation to 10°C while photosynthetic parameters , and I_k as well as activities of PePC and PePCk did not show any significant change after temperature acclimation. Contrary to P _max ^b , the activity of RuBPC was lower for algae acclimated to 3°-10°C, the observed response increasing with temperature. There was also a seasonal trend in the response of RuBPC, the ability to compensate for rapid temperature changes being higher in May. These results show that ice algae were photosynthetically adaptable in the range of temperatures tested. For RuBPC, adaptability developed seasonally when the environmental temperature started to fluctuate in May. Photosynthetic acclimatization to temperature may be of high ecological significance in extending the growth season of ice-algae.Contribution to the programs of GIROQ (Groupe interuniversitaire de recherches océanographiques du Québec) and of the Maurice-Lamontagne Institute (Department of Fisheries and Oceans)     

Nocturnal uptake and assimilation of nitrogen dioxide by C3 and CAM plants

In order to investigate nocturnal uptake and assimilation of NO2 by C3 and crassulacean acid metabolism (CAM) plants, they were fumigated with 4 microl l(-1) 15N-labeled nitrogen dioxide (NO2) for 8 h. The amount of NO2 and assimilation of NO2 by plants were determined by mass spectrometry and Kjeldahl-nitrogen based mass spectrometry, respectively. C3 plants such as kenaf (Hibiscus cannabinus), tobacco (Nicotiana tabacum) and ground cherry (Physalis alkekengi) showed a high uptake and assimilation during daytime as high as 1100 to 2700 ng N mg(-1) dry weight. While tobacco and ground cherry strongly reduced uptake and assimilation of NO2 during nighttime, kenaf kept high nocturnal uptake and assimilation of NO2 as high as about 1500 ng N mg(-1) dry weight. Stomatal conductance measurements indicated that there were no significant differences to account for the differences in the uptake of NO2 by tobacco and kenaf during nighttime. CAM plants such as Sedum sp., Kalanchoe blossfeldiana (kalanchoe) and Aloe arborescens exhibited nocturnal uptake and assimilation of NO2. However, the values of uptake and assimilation of NO2 both during daytime and nighttime was very low (at most about 500 ng N mg(-1) dry weight) as compared with those of above mentioned C3 plants. The present findings indicate that kenaf is an efficient phytoremediator of NO2 both during daytime and nighttime.     

Nitrogen uptake,assimilation and transport in barley in the presence of atmospheric nitrogen dioxide

Summary Exposure of the leaves of young barley plants to nitrogen dioxide (NO₂) was shown to affect the rate of translocation of N, the form in which it is transported in the xylem stream and the partitioning of N between roots and shoots. Following its entry through the leaves, NO₂ is assimilated by the plant into reduced nitrogenous compounds which accounted for the major increases in plant N content and growth. The various effects of atmospheric NO₂ upon barley seedlings were strongly influenced by nitrate supply to the roots.     

Alpine plants show species-level differences in the uptake of organic and inorganic nitrogen

As an estimate of species-level differences in the capacity to take up different forms of N, we measured plant uptake of ¹⁵N-NH₄ ⁺, ¹⁵N-NO₃ ^– and ¹⁵N, [1]-¹³C glycine within a set of herbaceous species collected from three alpine community types. Plants grown from cuttings in the greenhouse showed similar growth responses to the three forms of N but varied in the capacity to take up NH₄ ⁺, NO₃ ^– and glycine. Glycine uptake ranged from approximately 42% to greater than 100% of NH₄ ⁺ uptake; however, four out of nine species showed significantly greater uptake of either NH₄ ⁺ or NO₃ ^– than of glycine. Relative concentrations of exchangeable N at the sites of plant collection did not correspond with patterns of N uptake among species; instead, species from the same community varied widely in the capacity to take up NH₄ ⁺, NO₃ ^–, and glycine, suggesting the potential for differentiation among species in resource (N) use.     

Plant respiration in relation to growth, maintenance, ion uptake and nitrogen assimilation

Abstract Respiration in plants is generally observed to comprise two components: one proportional to the growth rate and the other to the plant dry mass. These components are usually interpreted as being related to the growth of new plant material and maintenance of existing plant material, respectively. By analysing data in this way, the respiratory costs of both structural synthesis and maintenance are observed to be greater in the root than the shoot. This contradicts current understanding of the biochemistry of the processes involved. The basic model is developed to incorporate three additional processes. The first is the cost of ion uptake for plant growth. The second allows for the fact that the site of nitrogen assimilation into amino acids may differ from the site of utilization for protein synthesis: when ammonium is supplied, this is incorporated immediately into amino acids owing to its toxicity to the plants; when nitrate is supplied it may be reduced either in the shoot or root, or both, and subsequently transported around the plant for utilization. The third process to be included is an energy cost for the uptake of ions to balance efflux from the root system. The theory is consistent with experimental observation and provides a means of understanding and interpreting respiration and nitrogen metabolism in plants.     

Leaf nitrogen dioxide uptake coupling apoplastic chemistry, carbon/sulfur assimilation, and plant nitrogen status

Emission and plant uptake of atmospheric nitrogen oxides (NO + NO₂) significantly influence regional climate change by regulating the oxidative chemistry of the lower atmosphere, species composition and the recycling of carbon and nutrients, etc. Plant uptake of nitrogen dioxide (NO₂) is concentration-dependent and species-specific, and covaries with environmental factors. An important factor determining NO₂ influx into leaves is the replenishment of the substomatal cavity. The apoplastic chemistry of the substomatal cavity plays crucial roles in NO₂ deposition rates and the tolerance to NO₂, involving the reactions between NO₂ and apoplastic antioxidants, NO₂-responsive germin-like proteins, apoplastic acidification, and nitrite-dependent NO synthesis, etc. Moreover, leaf apoplast is a favorable site for the colonization by microbes, which disturbs nitrogen metabolism of host plants. For most plant species, NO₂ assimilation in a leaf primarily depends on the nitrate (NO₃ ⁻) assimilation pathway. NO₂–N assimilation is coupled with carbon and sulfur (sulfate and SO₂) assimilation as indicated by the mutual needs for metabolic intermediates (or metabolites) and the NO₂-caused changes of key metabolic enzymes such as phosphoenolpyruvate carboxylase (PEPc) and adenosine 5′-phosphosulfate sulfotransferase, organic acids, and photorespiration. Moreover, arbuscular mycorrhizal (AM) colonization improves the tolerance of host plants to NO₂ by enhancing the efficiency of nutrient absorption and translocation and influencing foliar chemistry. Further progress is proposed to gain a better understanding of the coordination between NO₂–N, S and C assimilation, especially the investigation of metabolic checkpoints, and the effects of photorespiratory nitrogen cycle, diverse PEPc and the metabolites such as cysteine, O-acetylserine (OAS) and glutathione.     

The influence of root assimilated inorganic carbon on nitrogen acquisition/assimilation and carbon partitioning

Understanding of the influences of root-zone CO2 concentration on nitrogen (N) metabolism is limited. The influences of root-zone CO2 concentration on growth, N uptake, N metabolism and the partitioning of root assimilated 14C were determined in tomato (Lycopersicon esculentum). Root, but not leaf, nitrate reductase activity was increased in plants supplied with increased root-zone CO2. Root phosphoenolpyruvate carboxylase activity was lower with NO3(-)- than with NH4(+)-nutrition, and in the latter, was also suppressed by increased root-zone CO2. Increased growth rate in NO3(-)-fed plants with elevated root-zone CO2 concentrations was associated with transfer of root-derived organic acids to the shoot and conversion to carbohydrates. With NH4(+)-fed plants, growth and total N were not altered by elevated root-zone CO2 concentrations, although 14C partitioning to amino acid synthesis was increased. Effects of root-zone CO2 concentration on N uptake and metabolism over longer periods (> 1 d) were probably limited by feedback inhibition. Root-derived organic acids contributed to the carbon budget of the leaves through decarboxylation of the organic acids and photosynthetic refixation of released CO2.