Rates Of Ammonium Assimilation Research Articles

Nitrate and ammonium fluxes to diatoms and dinoflagellates at a single cell level in mixed field communities in the sea

Growth of large phytoplankton is considered to be diffusion limited at low nutrient concentrations, yet their constraints and contributions to carbon (C) and nitrogen fluxes in field plankton communities are poorly quantified under this condition. Using secondary ion mass spectrometry (SIMS), we quantified cell-specific assimilation rates of C, nitrate, and ammonium in summer communities of large phytoplankton when dissolved inorganic nitrogen concentrations are low in temperate coastal regions. Chain-forming diatoms composed 6% of total particulate organic carbon, but contributed 20% of C assimilation, 54% of nitrate assimilation and 32% of ammonium assimilation within the plankton community. In contrast, large dinoflagellates composed 11% of total POC, and contributed 14% of the C assimilation, 4% of ammonium and 9% of nitrate assimilation within the plankton community. Measured cell-specific C and nitrate assimilation rate match the Redfield ratio and the maximal nitrate assimilation in Chaetoceros spp. predicted by mass transfer theory. However, average ammonium assimilation rates were 30 and 340% higher than predicted by mass transfer theory in Tripos/Ceratium and Chaetoceros, respectively, suggesting that microbial interactions in the phycosphere may facilitate substantial luxury ammonium uptake by Chaetoceros in environments with fluctuating nitrate concentrations.

Mechanisms of nitrogen attenuation from seawater by two microbial mats

Microbial mats, due to their high microbial diversity, have the potential to express most biogeochemical cycling processes, highlighting their prospective use in bioremediation of various environmental contaminants. In this study the mechanisms of nitrogen attenuation were investigated in naturally occurring microbial mats from Elkhorn Slough, Monterey Bay, CA, USA, and Baja California Sur, Mexico. Key processes responsible for this removal were evaluated using quantification of functional genes related to nitrification, denitrification, and nitrogen fixation. Both microbial mats were capable of removing high (up to 2 mM) concentrations of ammonium and nitrate. Ammonium assimilation rates measured for Elkhorn Slough mats showed that this process was responsible for most of the ammonium uptake in these mats. While Elkhorn Slough mats did not show any evidence of nitrogen removal pathways other than microbial assimilation, Baja mats exhibited the potential for nitrification, denitrification, and DNRA as well as assimilation. The results of this study demonstrate the potential of microbial mats for bioremediation of nitrogenous pollutants independent of the mechanisms responsible for their removal.

Diazotroph community structure and the role of nitrogen fixation in the nitrogen cycle in the Chukchi Sea (western Arctic Ocean)

AbstractMarine nitrogen fixation occurs not only in subtropical and tropical regions but also in colder regions. However, the distribution of diazotrophs, nitrogen fixation rate, and its contribution to the nitrogen cycle in the Arctic Ocean remain poorly understood. We examined the diazotroph community structure and activity in the shelf and off‐shelf regions of the Chukchi Sea, western Arctic Ocean, during late summer 2015. The nitrate and ammonium assimilation rates were determined simultaneously to gain insights into the role of nitrogen fixation in the nitrogen cycle of the region. The diazotroph community determined by Illumina sequencing was mainly composed of Cluster III nifH phylotypes (putative anaerobes), accounting for > 80% of the total sequences examined, except for one surface sample. This result is strikingly different from previous findings in other oceanic regions. The nifH sequences other than those from Cluster III were mostly affiliated with UCYN‐A2 (symbiotic cyanobacteria), which accounted for < 15% of the total sequences. UCYN‐A2 tended to be abundant (maximum 2.9 × 103 copies L−1) in the high‐temperature low‐salinity water mass that is characteristic of Pacific‐originating water. Nitrogen fixation rate was detectable at most stations, with a range of 0.08–3.60 nmol L−1 d−1, displaying no clear relationship with depth (light intensity) or nitrate or ammonium concentration. Nitrogen fixation locally exceeded nitrate assimilation, but accounted for 1.00% at most in the total nitrogen assimilation in the euphotic zone.

High irradiance improves ammonium tolerance in wheat plants by increasing N assimilation

Ammonium is a paradoxical nutrient ion. Despite being a common intermediate in plant metabolism whose oxidation state eliminates the need for its reduction in the plant cell, as occurs with nitrate, it can also result in toxicity symptoms. Several authors have reported that carbon enrichment in the root zone enhances the synthesis of carbon skeletons and, accordingly, increases the capacity for ammonium assimilation. In this work, we examined the hypothesis that increasing the photosynthetic photon flux density is a way to increase plant ammonium tolerance. Wheat plants were grown in a hydroponic system with two different N sources (10mM nitrate or 10mM ammonium) and with two different light intensity conditions (300μmolphotonm−2s−1 and 700μmolphotonm−2s−1). The results show that, with respect to biomass yield, photosynthetic rate, shoot:root ratio and the root N isotopic signature, wheat behaves as a sensitive species to ammonium nutrition at the low light intensity, while at the high intensity, its tolerance is improved. This improvement is a consequence of a higher ammonium assimilation rate, as reflected by the higher amounts of amino acids and protein accumulated mainly in the roots, which was supported by higher tricarboxylic acid cycle activity. Glutamate dehydrogenase was a key root enzyme involved in the tolerance to ammonium, while glutamine synthetase activity was low and might not be enough for its assimilation.

Inorganic nitrogen assimilation and regeneration in the coastal upwelling region of the Iberian Peninsula

N‐regeneration and N‐assimilation rates were investigated in surface waters of the Iberian Peninsula, northeast Atlantic, during a period of seasonal upwelling. Stations were selected on the basis of near–real‐time remotely sensed data to provide a contrast between upwelling and oligotrophic conditions. Inorganic nitrogen (NO−3 and NH+4) was simultaneously regenerated and assimilated in the short term (i.e., hours), highlighting the importance of N‐regeneration in sustaining productivity. The regeneration of both NH+4 and NO−3 would introduce error in N‐assimilation rate estimations using 15N techniques, if not accounted for. The rate of ammonium assimilation (ρNH+4) frequently exceeded the rate of NO−3 assimilation (ρNO−3). Although the assimilation of NO−3 and NH+4 continued at night, dark NO−3 assimilation was lower than during the day. The separate oxidation steps of the nitrification process occurred at similar rates at most stations, and NO−2 oxidation made a significant, but variable, contribution to hourly phytoplankton NO−3 uptake. NO−2 accumulated in near‐surface waters of stations experiencing upwelling, suggesting an imbalance between production and consumption. At these stations, NO−2 release by phytoplankton may have stimulated NO−2 oxidation, uncoupling it from NH+4 oxidation. 15N‐based f‐ratio as an estimate of exportable production must be corrected for isotope dilution, and this requires information on NH+4 regeneration and nitrification rate; in addition, the assumed close coupling between the separate oxidation steps of nitrification must be verified. These corrections may be especially pertinent to upwelling regions, which represent an important route for new‐N input to the surface ocean.

Isotopic composition of particulate organic nitrogen and its relationship to nitrate assimilation in the Mediterranean Sea

SUMMARY: The shifts in the 15 N: 14 N ratio of particulate organic nitrogen (PON) suspended in the upper layer of the ocean have normally been interpreted in terms of changes in the isotopic composition of the source nitrogen used by the phytoplankton (nitrate, ammonium and dinitrogen gas) and/or the fractionation associated with the assimilation processes ( 14 N is preferentially taken up by phytoplankton). The objective of the present study was to test which process, isotopic fractionation during NO3 - uptake or reduced N-form consumption by phytoplankton, was more important in explaining the variations in the 15N signature of PON in the surface layer of a geostrophic front area (northwest Alboran Sea, Mediterranean Sea). Samples of PON were collected at 6 different stations during 4 seasonal surveys. The particulate organic carbon (POC) and PON concentrations averaged over the mixed layer (ML) ranged from 142 and 23 mg L-1 in summer to 104 and 13 mg L-1 in autumn respectively. The C:N molar ratio of organic matter also varied seasonally from 7.2 in summer, winter and spring to 9.3 in autumn. The range of variation of the d15N of PON in the ML was –0.1‰ to 6.5‰. The lowest values of d15N (lower than 1‰) were obtained in summer and winter. However, the d 15 N of PON in autumn and spring was higher than 3.5‰. Consequently, the particulate matter in the ML was enriched in 15N by about 1.5‰ in autumn and spring with respect to sum mer and winter. The d 15 N of PON in the surface layer was negatively correlated with nitrate concentration (r = -0.62, n = 22, p = 0.002). The d15N was also negatively correlated with the proportion of nitrate to total inorganic nitrogen (i.e. the sum of nitrate, nitrite and ammonium). Furthermore, there was a negative correlation between nitrate relative to nitrate plus ammonium assimilation rates (rNO 3 –) and the d15N (r = –0.71, n = 20; p<0.001) for the surface samples. rNO 3 – was particularly low (lower than 0.2) for the surface samples collected during autumn, when primary productivity mainly depended on the consumption of ammonium coming from the recycling of organic matter. Therefore, d15N of seston was a sensitive indicator of the transition from productivity based on nitrate to productivity based on regenerated ammonium.

Modelling nitrogen assimilation of Escherichia coli at low ammonium concentration

Modelling is an important methodology in systems biology research. In this paper, we presented a kinetic model for the complex ammonium assimilation regulation system of Escherichia coli. Based on a previously published model, the new model included AmtB mediated ammonium transport and AmtB regulation by GlnK. Protein concentrations and several parameter values were determined or refined based on new experimental data. Steady state analysis of the model showed that the expression of AmtB increased the ammonium assimilation rate 4–5-fold at external ammonium concentrations as low as 5 μM. Model analysis also suggested that AmtB and GS levels were coupled to maximize the assimilation flux and to avoid a possible negative ammonia diffusion flux. In addition, model simulation of the short term dynamic response to increased external ammonium concentrations implied that the maximal rate for GlnB/GlnK uridylylation/deuridylylation might be higher for a quick response to environmental changes.

METABOLIC AND ECOLOGICAL CONSTRAINTS IMPOSED BY SIMILAR RATES OF AMMONIUM AND NITRATE UPTAKE PER UNIT SURFACE AREA AT LOW SUBSTRATE CONCENTRATIONS IN MARINE PHYTOPLANKTON AND MACROALGAE1

Marine phytoplankton and macroalgae acquire important resources, such as inorganic nitrogen, from the surrounding seawater by uptake across their entire surface area. Rates of ammonium and nitrate uptake per unit surface area were remarkably similar for both marine phytoplankton and macroalgae at low external concentrations. At an external concentration of 1 μM, the mean rate of nitrogen uptake was 10±2 nmol·cm−2·h−1 (n=36). There was a strong negative relationship between log surface area:volume (SA:V) quotient and log nitrogen content per cm2 of surface (slope=−0.77), but a positive relationship between log SA:V and log maximum specific growth rate (μmax; slope=0.46). There was a strong negative relationship between log SA:V and log measured rate of ammonium assimilation per cm2 of surface, but the slope (−0.49) was steeper than that required to sustain μmax (−0.31). Calculated rates of ammonium assimilation required to sustain growth rates measured in natural populations were similar for both marine phytoplankton and macroalgae with an overall mean of 6.2±1.4 nmol·cm−2·h−1 (n=15). These values were similar to maximum rates of ammonium assimilation in phytoplankton with high SA:V, but the values for algae with low SA:V were substantially less than the maximum rate of ammonium assimilation. This suggests that the growth rates of both marine phytoplankton and macroalgae in nature are often constrained by rates of uptake and assimilation of nutrients per cm2 surface area.

Diurnal cycle and kinetics of ammonium assimilation in the green alga Ulva pertusa

The kinetics of ammonium assimilation was investigated in Ulva pertusa (Chlorophyceae, Ulvales) from northeastern New Zealand. Ammonium assimilation exhibited Michaelis–Menten kinetics with a maximum rate of assimilation (Vmax) of 54 ± 5 μmol g−1 dry weight h−1 and half-saturation constant (Km) of 23 ± 8 μM. In contrast, values for ammonium uptake were considerably higher with a Vmax of 316 ± 59 μmol g−1 dry weight h−1 and Km of 135 ± 46 μM. At environmentally relevant ammonium concentrations (5 μM), assimilation accounted for most (70%) of the ammonium taken up. Darkness decreased the maximum rate of ammonium assimilation by 83%. We investigated the hypothesis that rates of biosynthetic processes are greater in the early part of the day in Ulva. Consistent with this hypothesis, the maximum rate of ammonium assimilation in U. pertusa peaked in the morning and coincided with low levels of the photosynthetic product sucrose, which peaked in the afternoon. There was a diurnal cycle in the rate of ammonium uptake and assimilation in light and dark, but the amplitude was much greater for assimilation than uptake. Moreover, our data suggest that net ammonium assimilation only occurs during the day in U. pertusa. We suggest that two major roles for diurnal cycles are minimisation of interspecific competition for resources and metabolic costs.

Temporal variability in nitrification rates and related biogeochemical factors in Monterey Bay, California, USA

The scales of temporal variability in the rate of ammonium oxidation and a suite of related biogeochemical parameters were investigated in the surface layer of the water column of Monterey Bay, California, on the west coast of the USA. Samples were collected on bimonthly (approx. every 2 mo) cruises during the course of 2 yr. The signal of El Nino was evident in the water column temperature and nitrate distributions during the initial cruise in February 1998. Throughout the 2 yr period, variation in temperature and salinity was small and consistent with seasonal upwelling. The biological parameters, however, varied much more than might have been predicted by the minimal variability in hydrographic signals. Ammonium assimilation rates, chlorophyll a con- centrations and bacterial abundance varied by 41-, 65- and 147-fold, respectively. Ammonium oxida- tion rates often exhibited a subsurface maximum near the bottom of the euphotic zone, and varied by a factor of 4.5 over the 2 yr study. The presence of the rate maximum was not obviously related to the dominant seasonal signals of hydrography or to other biological variables. Nitrification rates were much less variable in the bay than in Elkhorn Slough, a small estuary that opens onto the bay. The small variability in ammonium oxidation rates may be related to more dynamic responses in the com- munity composition of the ammonia-oxidizing bacteria. Different groups of ammonia-oxidizing bac- teria were detected as dominants in clone libraries from the Monterey Bay and Elkhorn Slough. The time scale of bimonthly sampling was evidently inadequate to resolve the response of nitrification rates to direct physical forcing in the bay.

CONTRASTING EFFECTS OF METHIONINE SULFOXIMINE ON UPTAKE AND ASSIMILATION OF AMMONIUM IN ULVA INTESTINALIS (CHLOROPHYCEAE)1

Ammonium is assimilated in algae by the glutamine synthetase (GS)–glutamine:2‐oxoglutarate aminotransferase pathway. In addition to the assimilation of external ammonium taken up across the cell membrane, an alga may have to reassimilate ammonium derived from endogenous sources (i.e. nitrate reduction, photorespiration, and amino acid degradation). Methionine sulfoximine (MSX), an irreversible inhibitor of GS, completely inhibited GS activity in Ulva intestinalis L. after 12 h. However, assimilation of externally derived ammonium was completely inhibited after only 1–2 h in the presence of MSX and was followed by production of endogenous ammonium. However, endogenous ammonium production in U. intestinalis represented only a mean of 4% of total assimilation attributable to GS. The internally controlled rate of ammonium uptake (Vi) was almost completely inhibited in the presence of MSX, suggesting that Vi is a measure of the maximum rate of ammonium assimilation. After complete inhibition of ammonium assimilation in the presence of MSX, the initial or surge (Vs) rate of ammonium uptake in the presence of 400 μM ammonium chloride decreased by only 17%. However, the amount that the rate of ammonium uptake decreased by was very similar to the uninhibited rate of ammonium assimilation. In addition, the decrease in the rate of ammonium uptake in darkness (in the absence of MSX) in the presence of 400 μM ammonium chloride matched the decrease in the rate of ammonium assimilation. However, in the presence of 10 μM ammonium chloride, MSX completely inhibited ammonium assimilation but had no effect on the rate of uptake.

Nitrogen status and metabolism in the green seaweed Enteromorpha intestinalis: an examination of three natural populations

MEPS Marine Ecology Progress Series Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsTheme Sections MEPS 249:133-144 (2003) - doi:10.3354/meps249133 Nitrogen status and metabolism in the green seaweed Enteromorpha intestinalis: an examination of three natural populations Neill G. Barr*, T. Alwyn V. Rees Leigh Marine Laboratory and School of Environmental and Marine Sciences, University of Auckland, PO Box 349, Warkworth, New Zealand *Email: n.barr@auckland.ac.nz ABSTRACT: Nitrogen metabolism in Enteromorpha intestinalis from 3 contrasting populations in the Auckland region, New Zealand, was investigated. The sites (intertidal flats at Laingholm in the Manukau Harbour and Mangemangeroa and rockpools at Waterfall Reef) were chosen as they provided a range of ammonium enrichment. Seawater at Laingholm had higher levels of ammonium and total inorganic nitrogen compared with the other 2 sites. However, mean nitrate levels in Mangemangeroa seawater were similar to those measured at Laingholm. Seawater at Waterfall Reef had the lowest concentrations of ammonium and nitrate. At all 3 sites, E. intestinalis was found high in the intertidal. At Laingholm, the ammonium-enriched site, plants were immersed for 90 min at high tide; during this period the plants took up sufficient ammonium to increase their nitrogen content by 10%. The higher levels of inorganic nitrogen in the seawater at Laingholm resulted in higher levels of tissue nitrogen, chl a and b, glutamine and asparagine, and lower nitrogen-specific rates of ammonium assimilation (measured in the laboratory in the presence of saturating concentrations of ammonium) in E. intestinalis at this site. There were strong positive correlations between seawater concentrations of ammonium and the level of glutamine and between levels of chl a and b and tissue nitrogen in E. intestinalis. The nitrogen-specific rate of ammonium assimilation decreased as the nitrogen content of the plant increased and reached a minimum value above a nitrogen content of 1 to 2% or a glutamine level of 5 µmol per g dry weight (DW). This glutamine level corresponded to a seawater ammonium concentration of 6 µM. The nitrogen-specific rate of ammonium assimilation provides a novel and important bioindicator of the link between the concentration of inorganic nitrogen in the environment and the nitrogen status of the plant. KEY WORDS: Amino acids · Ammonium assimilation · Bioindicators · Chlorophyll · Enteromorpha intestinalis · Nitrogen status Full text in pdf format PreviousNextExport citation RSS - Facebook - Tweet - linkedIn Cited by Published in MEPS Vol. 249. Online publication date: March 10, 2003 Print ISSN: 0171-8630; Online ISSN: 1616-1599 Copyright © 2003 Inter-Research.

KINETICS OF AMMONIUM ASSIMILATION IN TWO SEAWEEDS, ENTEROMORPHA SP. (CHLOROPHYCEAE) AND OSMUNDARIA COLENSOI (RHODOPHYCEAE)

The kinetics of ammonium assimilation were investigated in two seaweeds from northeastern New Zealand, Enteromorpha sp. (Chlorophyceae, Ulvales) and Osmundaria colensoi (Hook. f. et Harvey) R.E. Norris (Rhodophyceae, Ceramiales), with the use of a recently developed method for measuring assimilation. In contrast to ammonium uptake, which was nonsaturable, ammonium assimilation exhibited Michaelis–Menten kinetics in both species. Maximum rates of assimilation (Vmax) were 27 and 12 μmol·(g DW)−1·h−1 for Enteromorpha sp. and O. colensoi, respectively, with half‐saturation (Km) constants for assimilation of 18 and 41 μM. At environmentally relevant concentrations, assimilation accounted for all of the ammonium taken up by both species. The maximum rate of assimilation in Enteromorpha sp. resembled very closely that of the ammonium assimilatory enzyme, glutamine synthetase, when activities of the latter were measured in the presence of subsaturating substrate (glutamate and ATP) concentrations. Moreover, the initial rate of glutamine production (measured with HPLC) following ammonium enrichment was almost identical to the rates determined above. The rate of ammonium assimilation was therefore determined by three independent methods, two of which involve in vivo measurements, and it is suggested that the use of assimilation kinetics may be useful when examining the nutrient relations of seaweeds.

Allometric evidence for the dominant role of surface cells in ammonium metabolism and photosynthesis in northeastern New Zealand seaweeds

Relationships between seaweed morphology and the metabolism of ammonium and carbon were examined for 9 seaweeds from northeastern New Zealand. Maximum rates of surge ammonium uptake and ammonium assimilation, in addition to the maximum ammonium storage pool, were strongly positively correlated with seaweed surface area:volume (SA:V) ratio when expressed per g dry weight, but were largely independent of SA:V ratio when expressed per cm 2 surface area. This suggests that ammomum metabolism may be confined largely to the outermost cell layers. Similar scaling properties were found for rates of photosynthesis and, to d lesser extent, respiration. Rates of photosynthesis were similar to those derived from the literature for northern hemisphere seaweeds, while respiration rates were slightly lower among New Zealand seaweeds. These comparatively minor geographic differences suggest that previously documented differences in rates of ammonium uptake between the 2 regions are not related to carbon metabolism. Maximum rates of ammonium assimilation were determined by measuring the internally controlled rate of ammonium uptake, which closely matched the assimilation rate in the green seaweed Ulva sp., the red seaweed Osmundaria colensoi, and the brown seaweed Zonaria turneriana.

Sucrose-feeding leads to increased rates of nitrate assimilation, increased rates of α-oxoglutarate synthesis, and increased synthesis of a wide spectrum of amino acids in tobacco leaves

To investigate the importance of the sugar supply for the regulation of nitrogen and organic acid metabolism, various sugars and nitrogenous compounds were supplied for 8 h to detached tobacco leaves in low light. (i) In control leaves supplied with water, there was a large decrease of the Nia transcript level, a 50% decline of nitrate reductase (NR) activity, starch increased and sugars remained low, nitrate decreased by 50%, and amino acids increased only slightly during the 8 h incubation. About half of the nitrogen accumulating in amino acids was present in glutamine (Gln). (ii) When 25 mM sucrose was supplied, the in-vivo rate of nitrate assimilation (estimated from the accumulation of ammonium and amino acids) increased 2-fold. The Nia transcript level still decreased, but the decline of NR activity was less pronounced and NR activation was increased. The in-vivo net rate of ammonium assimilation (estimated from the accumulation of amino acids) also doubled after feeding sucrose. Ammonium and glutamate (Glu) decreased and Gln rose markedly, showing that in-vivo activity of glutamine synthetase had been stimulated. Glutamine still accounted for about half of the nitrogen, indicating that sucrose does not selectively stimulate glutamine synthase. Glutamate and aspartate decreased and all the minor amino acids increased, showing that the amino acid biosynthesis pathways are activated by sucrose. There was a decrease of 3-phosphoglycerate (3PGA) and phosphoenolpyruvate (PEP) and a large increase of α-oxoglutarate, showing that the flow of carbon from glycolysis into organic acids has been stimulated by sucrose. (iii) The changes of 3PGA, PEP, α-oxoglutarate, Glu, aspartate and the minor amino acids were smaller when 50 mM glucose was supplied, even though the internal levels of sugars at the end of the incubation resembled those found after feeding 25 mM sucrose. This indicates that the signals that regulate nitrogen and respiratory metabolism are derived from the uptake or metabolism of sucrose, rather than glucose. (iv) A different spectrum of changes was found when 20 mM nitrate was supplied. The estimated rate of nitrate assimilation increased 2-fold, and this was accompanied by an increase of NR activity but not of NR activation. Nitrate-feeding did not lead to a decrease of Glu, and the increase of minor amino acids was slightly smaller than with sucrose. There was a decrease of sugars, starch, and hexose phosphates, but 3PGA and PEP were not significantly decreased and isocitrate increased instead of α-oxoglutarate. (v) A different spectrum of changes was also found when 10 mM Gln was supplied. The estimated rate of nitrate assimilation decreased, and this was accompanied by a decrease of NR activity and NR activation. Glutamate did not decrease, and the increase of minor amino acids was smaller than with sucrose. Starch and sugars remained high and, although hexose phosphates decreased, 3PGA and PEP were not significantly decreased. Isocitrate remained unaltered and the increase of α-oxoglutarate was smaller than after supplying sucrose. (vi) When 25 mM sucrose was added together with 20 mM nitrate or 10 mM Gln, the effect on NR activity, NR activation and the estimated rate of nitrate assimilation was additive to the effect of nitrate, and antagonistic to the effect of Gln. Sucrose still led to a decrease of Glu, an increase of the minor amino acids, a decrease of 3PGA and PEP, and an increase of α-oxoglutarate when it was supplied together with nitrate or Gln. (vii) It is concluded that sucrose initiates a co-ordinate activation of nitrate assimilation, ammonium assimilation, amino acid biosynthesis, and α-oxoglutarate synthesis. Sucrose acts in concert with nitrate and antagonistically to Gln to increase NR activity and nitrate assimilation, and complements the action of nitrate and Gln to increase the flow of nitrogen from ammonium into amino acids, and to increase α-oxoglutarate formation.

MEASURING RATES OF AMMONIUM ASSIMILATION IN MARINE ALGAE: USE OF THE PROTONOPHORE CARBONYL CYANIDE m‐CHLOROPHENYLHYDRAZONE TO DISTINGUISH BETWEEN UPTAKE AND ASSIMILATION

A method for determining rates of ammonium assimilation in marine algae is described. Ammonium assimilation is defined as the decrease in total (medium + cellular) ammonium. The protonophore carbonyl cyanide m‐chlorophenylhydrazone (CCCP) was used to distinguish between uptake and assimilation of ammonium. Ammonium uptake by nitrogen‐replete and nitrogen‐starved cells of the diatom Phaeodactylum tricornutum Bohlin and the green macroalga Enteromorpha sp. was completely (98%–99%) inhibited in the presence of 100 μM CCCP. In addition to inhibiting further uptake of ammonium, CCCP promoted the release of unassimilated ammonium by nitrogen‐replete and nitrogen‐starved P. tricornutum and Enteromorpha that had been allowed to take up ammonium for a period. Most (97.5%) of preaccumulated 14C‐methylammonium was released by nitrogen‐starved P. tricornutum in the presence of CCCP. Specific rates of ammonium assimilation in nitrogen‐replete cultures of P. tricornutum were identical to the maximum growth rate, but specific rates in nitrogen‐starved cultures were fourfold greater. Rates of ammonium assimilation in Enteromorpha during both the surge and the internally controlled uptake phases were the same as the internally controlled rate of uptake, suggesting that the latter is a reliable measure of the maximum rate of assimilation.

EFFECT OF MALTOSE RELEASE ON UPTAKE AND ASSIMILATION OF AMMONIUM BY SYMBIOTIC CHLORELLA (CHLOROPHYTA)1

ABSTRACTWhen incubated at pH 4–5, Chlorella freshly isolated from symbiosis with Hydra viridissima PALLAS 1766 (green hydra) release large amounts of photosynthetically fixed carbon in the form of maltose, and assimilation of inorganic N is inhibited. Physiological responses to N starvation of the cultured 3N813A strain of maltose‐releasing Chlorella differed from those caused by 48 h of maltose release induced by low pH. N starvation increased rates of ammonium assimilation at pH 7.0 in light or darkness, and ammonium assimilation in darkness stimulated cell respiration. In contrast, cells pretreated at pH 5.0 to induce maltose release were unable to take up ammonium at pH 7.0 unless supplied with an external carbon source such as bicarbonate, acetate, or succinate, and rates of uptake were similar to control cells. Freshly isolated symbionts displayed a similar dependency. Rates of ammonium uptake by cells pretreated at pH 5.0 were reduced in darkness and did not stimulate cell respiration. N‐starved cells supplied with ammonium also showed a large short‐term increase in glutamine pools at the expense of glutamate, as might be expected if large amounts of ammonium were rapidly assimilated via glutamine synthetase/glutamate synthase, whereas after long‐term maltose release cells showed only a small increase in glutamine when supplied with ammonium. Furthermore, maltose release caused a fall in pool sizes of a number of amino acids, including glutamine and glutamate, and also caused a decrease in pool sizes of 2‐oxoglutarate and phospho‐enol‐pyruvate, which are required for ammonium assimilation into amino acids.Cells stimulated to synthesize and release maltose may be unable to assimilate ammonium and synthesize amino acids because of diversion of fixed carbon from N metabolism. We estimate that 40–50% affixed C is required for maximal maltose synthesis, whereas up to 30% fixed C is required for ammonium assimilation. These results are discussed in the context of host regulation of symbiotic algal growth.

Changes in intracellular amino acids and organic acids induced by nitrogen starvation and nitrate or ammonium resupply in the cyanobacterium Phormidium laminosum.

In Phormidium laminosum cells, nitrogen starvation caused a decrease in the intracellular levels of all amino acids, except glutamate, and an increase in the total level of the analyzed organic acids. The addition of nitrate or ammonium to N-starved cells resulted in substantial increases in the pool size of most amino acids. Upon addition of ammonium the total level of organic acids diminished, whereas it increased upon addition of nitrate, after a transient decay during the first minutes. Nitrogen resupply stimulated amino acid synthesis, the effect being faster and higher when ammonium was assimilated. The data indicate that nitrate and ammonium assimilation induced an enhancement of carbon flow through the glycolytic and the tricarboxylic-acid pathways to amino acid biosynthesis, with a concurrent decrease in the carbohydrate reserves. The results suggest that the availability of carbon skeletons limited the rate of ammonium assimilation, whereas the availability of reducing equivalents limited the rate of nitrate assimilation.

Estimating nitrogen transformation rates in surface aerobic soil of a paddy field

Rates of nitrogen transformations in the surface aerobic soil of a paddy field (fine-textured strong-gley soils) were estimated by combination of a mathematical model and parallel incubation experiments using 15N as a tracer. The mineralization rate was nearly constant throughout the incubation. However, the ammonium assimilation rate during the initial phase of the incubation was two to three times higher than the mineralization rate, and rapidly decreased with time. This decrease did not follow first-order kinetics for the ammonium content in soil. The nitrification rate increased slightly with time. Reduction of nitrate to ammonium was detectable in all the experiments, but its rate was very low (<50μg N kg h −1). The relative magnitudes of the rates of N transformations changed with the incubation temperature. The rates of denitrification plus nitrate assimilation were greatly enhanced at 30°C. The mineralization rate also increased with temperature, but the nitrification rate decreased slightly. The rates of N transformations in the surface layer, especially under aerobic condition, were from 10 to 100 times greater than those in the subsurface layer under anaerobic condition. This indicates that N transformations in the surface layers of paddy soils are quantitatively of great importance.

Partitioning of inorganic nitrogen assimilation between the roots and shoots of cerrado and forest trees of contrasting plant communities of South East Brasil.

Woody plants growing in cerrado and forest communities of south-east Brasil were found to have low levels of nitrate reductase activity in their leaves suggesting that nitrate ions are not an important nitrogen source in these communities. Only in the leaves of species growing in areas of disturbance, such as gaps and forest margins, were high levels of nitrate reductase present. When pot-grown plants were supplied with nitrate, leaves and roots of almost all species responded by inducing increased levels of nitrate reductase. Pioneer or colonizing species exhibited highest levels of nitrate reductase and high shoot: root nitrate reductase activities. Glutamine synthetase, glutamate synthase and glutamate dehydrogenase were present in leaves and roots of the species examined.15N-labelled nitrate and ammonium were used to compare the assimilatory characteristics of two species:Enterolobium contortisiliquum, with a high capacity to reduce nitrate, andCalophyllum brasiliense, of low capacity. The rate of nitrate assimilation in the former was five times that of the latter. Both species had similar rates of ammonium assimilation. Results for eight species of contrasting habitats showed that leaf nitrogen content increased in parallel with xylem sap nitrogen concentrations, suggesting that the ability of the root system to acquire, assimilate or export nitrate determines shoot nitrogen status. These results emphasise the importance of nitrogen transport and metabolism in roots as determinants of whole plant nitrogen status.