Two different isoforms of glucose-6-phosphate dehydrogenase (Glc6PDH; EC 1.1.1.49) have been partially purified from barley (Hordeum vulgare L., cv. Alfeo) roots. The procedure included an ammonium sulfate step, Q-Sepharose and Reactive Blue agarose chromatography, and led to 60-fold and 150-fold purification for the two enzymes, respectively. The Glc6PDH 1 isoform accounts for 17% of total activity of the enzyme in roots, and is very sensitive to the effects of NADP+/NADPH ratio and dithiothreitol; the Glc6PDH 2 isoform is less affected by reducing power and represents 83% of the total activity. The isoforms showed distinct pH optima, isoelectric points, Km for glucose-6-phosphate and a different electrophoretic mobility. The kinetic properties for the two enzymes were affected by ATP and metabolites. Both enzymes are inhibited to different extents by ATP when magnesium is omitted from the assay mixture, whereas the addition of ATP-Mg2+ had no effect on Glc6PDH activities. The Glc6PDH isoforms are usually present in the plastids and cytosol of plant cells. To verify the intracellular locations of the enzymes purified from barley roots, Glc6PDH was purified from isolated barley root plastids; this isoform showed kinetic parameters coincident with those found for Glc6PDH 1, suggesting a plastid location; the enzyme purified from the soluble fraction had kinetic parameters resembling those of Glc6PDH 2, confirming that this isoform is present in the cytosol of barley roots.

In barley (Hordeum vulgare L. var. Nure), glutamate synthesis and the production of reducing power by the oxidative pentose phosphate pathway (OPPP) are strictly correlated biochemical processes. NADH-GOGAT was the major root isoform, whose activity increased on a medium supplied with NH4+ or NO3-; by contrast, no noticeable variations could be observed in the leaves of plants supplied with nitrogen. In the leaves, the major isoform is Fd-GOGAT, whose activity increased under nitrogen feeding. G6PDH activity increased in the roots supplied with nitrogen; no variations were observed in the leaves. Moreover, an increase of the P2 isoform in the roots was measured, giving 13.6% G6PDH activity localized in the plastids under ammonium, and 25.2% under nitrate feeding conditions. Western blots confirmed that P2-G6PDH protein was induced in the roots by nitrogen. P1-G6PDH protein was absent in the roots and increased in the leaves by nitrogen supply to the plants. The changes measured in cytosolic G6PDH seem correlated to more general cell growth processes, and do not appear to be directly involved in glutamate synthesis. The effects of light on Fd-GOGAT is discussed, together with the possibility for P2-G6PDH to sustain nitrogen assimilation upon illumination.

Plants respond to changes of nutrient availability in the soil by modulating their root system developmental plan. This response is mediated by systemic changes of the nutritional status and/or by local perception of specific signals. The effect of nitrate on Arabidopsis (Arabidopsis thaliana) root development represents a paradigm of these responses, and nitrate transporters are involved both in local and systemic control. Ammonium (NH 4 + ) represents an important nitrogen (N) source for plants, although toxicity symptoms are often associated with high NH 4 + concentration when this is present as the only N source. The reason for these effects is still controversial, and mechanisms associating ammonium supply and plant developmental programs are completely unknown. We determined in Lotus japonicus the range of ammonium concentration that significantly inhibits the elongation of primary and lateral roots without affecting the biomass of the shoot. The comparison of the growth phenotypes in different N conditions indicated the specificity of the ammonium effect, suggesting that this was not mediated by assimilatory negative feedback mechanisms. In the range of inhibitory NH 4 + conditions, only the LjAMT1;3 gene, among the members of the LjAMT1 family, showed a strong increased transcription that was reflected by an enlarged topology of expression. Remarkably, the short-root phenotype was phenocopied in transgenic lines by LjAMT1;3 overexpression independently of ammonium supply, and the same phenotype was not induced by another AMT1 member. These data describe a new plant mechanism to cope with environmental changes, giving preliminary information on putative actors involved in this specific ammonium-induced response.

In plant cells, ammonium is considered the most convenient nitrogen source for cell metabolism. However, despite ammonium being the preferred N form for microalgae, at higher concentrations, it can be toxic, and can cause growth inhibition. Microalgae’s tolerance to ammonium depends on the species, with various taxa showing different thresholds of tolerability and symptoms of toxicity. In the environment, ammonium at high concentrations represents a dangerous pollutant. It can affect water quality, causing numerous environmental problems, including eutrophication of downstream waters. For this reason, it is important to treat wastewater and remove nutrients before discharging it into rivers, lakes, or seas. A valid and sustainable alternative to conventional treatments could be provided by microalgae, coupling the nutrient removal from wastewater with the production of valuable biomass. This review is focused on ammonium and its importance in algal nutrition, but also on its problematic presence in aquatic systems such as wastewaters. The aim of this work is to provide recent information on the exploitation of microalgae in ammonium removal and the role of ammonium in microalgae metabolism.

Summary• Temperature effects on growth, photosynthesis, respiration and nitrate reductase (NR) were studied in the cryophilic algae Koliella antarctica and ' Chlorella ' saccharophila , and in the mesophilic Chlorella sorokiniana .• Growth rate was measured as increase in optical density. Photosynthesis at saturating light and respiration in darkness were measured as O 2 exchange. NADH : NR was assayed in crude extracts.• The two cryophilic algae grew below 15 ° C, and C. sorokiniana above 20 ° C. Photosynthetic and respiration rates of K. antarctica and ' C .' saccharophila were elevated at 5 ° C, and peaked at 30 ° C. Arrhenius plots from 5 to 25 ° C were linear in K. antarctica , whereas in ' C .' saccharophila and C. sorokiniana they exhibited breaks at 15 and 20 ° C, respectively. Values for activation energy ( E a ) and the factor by which the rate increases with raising the temperature 10 ° C ( Q 10 ) differed. Nitrate reductase had its optimum at 25 ° C in cryophilic algae and at 35 ° C in C. sorokiniana .• We conclude that growth of cryophilic algae at low temperature is favoured by elevated photosynthesis and respiration rates, but that it could be limited by a high respiration : photosynthesis ratio.

The heavy metal pollution in soils and aquatic environments is a serious ecological problem. In the green-microalga Chlorella sorokiniana 211-8K (Chlorophyceae) exposed to ions of lead (Pb) and cadmium (Cd) we studied the metabolic responses to the toxicity of these two heavy metals. Our data indicate that both the pollutants alter the alga cell ultrastructure and its physiological characteristics (growth, photosynthesis, respiration, enzyme activities). The toxic effects of the two metals resulted time-dependent to the exposure. After 24 h of treatment with 250 μM Pb or Cd, photosynthesis was inhibited until to 77 and 86%, however respiration was strongly enhanced up to 300 and 350%, respectively. In the algal cells Pb or Cd exposure induced a reduction in the content of the total chlorophylls and a decrease of the soluble protein levels, significantly compromising the growth, particularly in cultures cadmium-treated. We report data on ultrastructural changes induced by the two heavy metals; they affected overall chloroplast ultrastructure of the alga. Most importantly, the O-acetyl-L-serine(thiol)lyase (OASTL) activity was appreciably increased after only 2 h of Cd exposure, indicating the existence of a link between the metal contamination and cysteine synthesis. Then, Chlorella sorokiniana cells seem to better tolerate high concentrations of Pb while appear to be more sensitive to Cd ions. These results provide some additional information that can lead to better understand consequences of heavy metal poisoning in microalgae.

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