Maize (Zea mays L.) plants were grown to the nine-leaf stage. Despite a saturating N supply, the youngest mature leaves (seventh position on the stem) contained little NO3- reserve. Droughted plants (deprived of nutrient solution) showed changes in foliar enzyme activities, mRNA accumulation, photosynthesis, and carbohydrate and amino acid contents. Total leaf water potential and CO2 assimilation rates, measured 3 h into the photoperiod, decreased 3 d after the onset of drought. Starch, glucose, fructose, and amino acids, but not sucrose (Suc), accumulated in the leaves of droughted plants. Maximal extractable phosphoenolpyruvate carboxylase activities increased slightly during water deficit, whereas the sensitivity of this enzyme to the inhibitor malate decreased. Maximal extractable Suc phosphate synthase activities decreased as a result of water stress, and there was an increase in the sensitivity to the inhibitor orthophosphate. A correlation between maximal extractable foliar nitrate reductase (NR) activity and the rate of CO2 assimilation was observed. The NR activation state and maximal extractable NR activity declined rapidly in response to drought. Photosynthesis and NR activity recovered rapidly when nutrient solution was restored at this point. The decrease in maximal extractable NR activity was accompanied by a decrease in NR transcripts, whereas Suc phosphate synthase and phosphoenolpyruvate carboxylase mRNAs were much less affected. The coordination of N and C metabolism is retained during drought conditions via modulation of the activities of Suc phosphate synthase and NR commensurate with the prevailing rate of photosynthesis.

Tobacco (Nicotiana tabacum L.) plants were subjected to a prolonged period of sulfur-deprivation to characterize molecular and metabolic mechanisms that permit control of primary N-metabolism under these conditions. Prior to the appearance of chlorotic lesions, sulfur-deprived tobacco leaves showed a strong decrease in the sulfate content and changes in foliar enzyme activities, mRNA accumulation and amino-acid pools. The basic amino acids glutamine, asparagine and arginine accumulated in the leaves of sulfur-deprived plants, while the foliar concentrations of aspartate, glutamate, serine or alanine remained fairly unchanged. Maximal extractable nitrate reductase (NR; EC 1.6.6.1) activity decreased strongly in response to sulfur-deprivation. The decrease in maximal extractable NR activity was accompanied by a decline in NR transcripts while the mRNAs of the plastidic glutamine synthetase (EC 6.1.3.2) or the beta-subunit of the mitochondrial ATP synthase were much less affected. Nitrate first accumulated in leaves of tobacco during sulfur-deprivation but then declined. An appreciable amount of nitrate was, however, present in severely sulfur-depleted leaves. The repression of NR gene expression is, therefore, not related to the decrease in the leaf nitrate level. However, glutamine- and/or asparagine-mediated repression of NR gene transcription is a possible mechanism of control in situations when glutamine and asparagine accumulate in leaves and provides a feasible explanation for the reduction in NR activity during sulfur-deprivation. The removal of reduced nitrogen from primary metabolism by redirection and storage as arginine, asparagine or glutamine combined with the down-regulation of nitrate reduction via glutamine- and/or asparagine-mediated repression of NR gene transcription may contribute to maintaining a normal N/S balance during sulfur-deprivation and indicate that the co-ordination of N- and S-metabolism is retained under these conditions.

The impact of increased plastidic glutamine synthetase (GS-2; EC 6.1. 3.2) activity on foliar amino-acid levels and on biomass production was examined in transgenic tobacco. For that, tobacco was transformed via Agrobacterium tumefaciens with a binary vector containing a tobacco GS-2 cDNA downstream of the leaf-specific soybean ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit gene promotor. Two transgenic tobacco lines with 15- to 18-fold higher foliar GS-2 transcript levels than the wild type were obtained. The GS-2 protein pools and the specific GS-2 activities were, however, only 2- to 2.3-fold higher in the leaves of the transgenic plants than in the leaves of the wild type. This discrepancy may reflect a post-transcriptional control of GS-2 protein accumulation. The increased GS-2 activity was correlated with a decrease in the leaf ammonium pool (3.7-fold) and an increase in the levels of some free amino acids, including glutamate (2. 5-fold) and glutamine (2.3-fold). The accumulation of soluble protein per unit fresh weight, however, remained unchanged. This result indicates that a process downstream of the synthesis of the primary organic products of N-assimilation is limiting leaf protein accumulation. Nevertheless, the overexpression of GS-2 stimulated the growth rate of the transgenic tobacco seedlings which, consequently, were larger (20-30% on a fresh-weight basis) than wild-type seedlings grown under identical conditions. This result suggests that GS-2 is the rate-limiting enzyme during biomass production in tobacco seedlings. The requirement for glutamate as the ammonium acceptor in the reaction catalysed by GS-2 may imply that there is co-regulation of GS-2 and ferredoxin dependent glutamate synthase (Fd-GOGAT; EC 1.4.7.1) gene expression. Increased leaf GS-2 activity had, however, no influence on the foliar Fd-GOGAT protein abundance. This result suggests that in tobacco leaves, more Fd-GOGAT is present than required to meet the demands of primary ammonium assimilation and that there is no strong interdependence between GS-2 and Fd-GOGAT protein expression.

Tomato (Lycopersicon esculentum L.) responded to a prolonged period of water stress with stomatal closure followed by premature flowering and the subsequent production of small fruits containing fertile seeds. Water stress was correlated with a net loss of protein from tomato leaves and the concomitant accumulation of free amino acids, reflecting the remobilization of leaf nitrogen to meet the N‐requirement for the rapid development of reproductive organs. We show by northern blot analysis of the transcript pools, and by immunoblot analysis of the protein levels that water stress stimulates tomato cytosolic glutamine synthetase (EC 6.1.3.2; GS‐1) gene expression, while plastidic glutamine synthetase (GS‐2) gene expression remains unchanged during drought. These results suggest a role of GS‐1 in the generation of glutamine for the transport of the nitrogen that is remobilized in tomato leaves in response to chronic water stress. The remobilization of leaf N during water stress appears to be. at least in part, initiated by a specific down‐regulation of the leaf transcript pool corresponding to the small subunit of ribulose‐1,5‐bisphosphate carboxylase/oxygenase.

A role for cytosohc glutamine sytithetase in the remobilization of leaf nittogen during water stress in tomato. -Physiol. Plant. 99: 241-248, Tomato (Lycopersicon esculentum L.) responded to a prolonged period of water stress with stomatal elosure followed by premature flowering and the subsequent production of small fruits containing fertile seeds. Water stress was correlated with a net loss of protein from tomato leaves and the concomitant aecumtilation of free amino acids, reflecting the remobilization of leaf nitrogen to meet the N-requirement for the rapid development of reproductive organs. We show by northem blot analysis of the transcript pools, and by immunobiot analysis ofthe protein levels that water stress stimulates tomato cytosolic glutamine synthetase (EC 6. L3.2; GS-1) gene expression, while plastidic gluiamine synthetase (GS-2) gene expression remains unchanged during drought. These results suggest a role of GS-1 in the generation of glutamine for the transport of the nitrogen that is remobilized in tomato leaves in response to chronic water stress. The remobilization of leaf N during water stress appears to be. at least in part, initiated by a speeifie dowti-regulation of the leaf transcript pool corresponding to the small subunit of ribulose-L5-bisphosphate earboxylase/oxygenase.