Overexpression of a Medicago truncatula stress-associated protein gene (MtSAP1) leads to nitric oxide accumulation and confers osmotic and salt stress tolerance in transgenic tobacco

The impact of Medicago truncatula stress-associated protein gene (MtSAP1) overexpression has been investigated in Nicotiana tabacum transgenic seedlings. Under optimal conditions, transgenic lines overexpressing MtSAP1 revealed better plant development and higher chlorophyll content as compared to wild type seedlings. Interestingly, transgenic lines showed a stronger accumulation of nitric oxide (NO), a signaling molecule involved in growth and development processes. This NO production seemed to be partially nitrate reductase dependent. Due to the fact that NO has been also reported to play a role in tolerance acquisition of plants to abiotic stresses, the responses of MtSAP1 overexpressors to osmotic and salt stress have been studied. Compared to the wild type, transgenic lines were less affected in their growth and development. Moreover, NO content in MtSAP1 overexpressors was always higher than that detected in wild seedlings under stress conditions. It seems that this better tolerance induced by MtSAP1 overexpression could be associated with this higher NO production that would enable seedlings to reach a high protection level to prepare them to cope with abiotic stresses.     

Stage of development is an important determinant in the effect of nitrate on photoassimilate (13C) partitioning in chicory (Cichorium intybus)

The effect of nitrogen supply to chicory plants on carbon partitioningbetween shoot, root and tuberized root was studied at differentstages of vegetative growth, using long-term ¹³CO₂ labelling-chaseexperiments. This approach was complemented by measurement ofstorage carbohydrates and activities of enzymes involved inroot sucrose metabolism (sucrose-sucrose fructosyl transferase(SST), sucrose synthase, invertase). In both young and matureplants, low resulted in a 30–35% decrease in ¹³C assimilation. However, the partitioningof ¹³C between shoot and root was affected differently at differentstages of development. In young plants, in which carbohydrateswere being used for structural root and shoot growth, neither¹³C shoot/¹³C root ratio nor root activities of the above enzymeswere modified by supply. In contrast, in mature plants storing large amounts of carbohydratesas fructan in the tuberized root, low caused the ratio to decrease from 0.6 to 0.2, despiteunchanged net flux of ¹³C from shoot to root. The extractableactivity of SST was elevated in mature plants, compared to youngplants, at both low and high , consistent with its role in fructan synthesis. However, matureplants grown at low exhibited SST activity double that of plants grown at high . From these results, it is concluded that the observeddecrease in shoot/root dry weight ratio at low supply is caused by increased utilization of carbohydratesfor storage due to elevated root SST activity. Key words: Chicory, nitrate, ¹³C, shoot/root ratio, fructans, SST     

Phosphate availability in combination with nitrate availability affects root yield and chicon yield and quality of Belgian endive (Cichorium intybus)

This study investigated the effects of nitrate and phosphate nutrition on chicory tap root development and chicon quality. Plants of chicory (Cichorium intybus flash) were grown on four concentrations of nitrate and phosphate: 3 mM NO₃ / 1 mM PO ₄ ^3– , high N and high P (control plants, N / P); 3 mM NO ₃ ^– / 0.05 mM H₂PO^3– ₄, high N and low P (N / p); 0.6 mM NO₃ / 1 mM PO ₄ ^3– , low N and high P (n / P); 0.6 mM NO ₃ ^– / 0.05 mM PO ₄ ^3– , low N and low P (n / p). The results suggested that, nitrogen limitation had the greatest impact on the shoot/root dry weight ratio. Only small changes in the shoot/root dry weight could be attributed to P limitation alone. Compared with the control, N limitation caused a marked increase in root SST activity (sucrose sucrose fructosyl transferase, the enzyme responsible for fructan synthesis in roots), the effect of P limitation on SST activity was less pronounced. The activity of SS (sucrose synthase) was also noticeably elevated at the early sample data by N limitation. N and P uptake were estimated by the amount of N and P accumulated by the whole plant during the vegetative period. With N limitation, P accumulation was decreased by 40-60% over the experimental period. The effects of P limitation on N accumulation were more variable, N uptake was 60% lower than the control during the tuberizing period (107 days after sowing). With N limitation, P concentrations in roots were lowered by 20-25%. With P limitation, total N concentration in roots decreased by 50% relative to the control, while nitrate concentration was increased more than 8 fold. These effects were detected only at 107 DAS. The amino acid content of roots was not affected by P limitation, however, N limitation altered strongly total amino acids. P limitation did alter the relative amino acid composition of roots early in the vegetative period: Roots harvested at the end of vegetative period were forced in the dark to produce an etiolated bud, the edible chicon. High N and high P fertility (N/P) were associated to a poor chicon yield and quality. However the presence of low P during vegetative growth moderates adverse effects of high nitrate and greatly improved chicon yeild and quality.     

Concerted modulation of alanine and glutamate metabolism in young Medicago truncatula seedlings under hypoxic stress

The modulation of primary nitrogen metabolism by hypoxic stress was studied in young Medicago truncatula seedlings. Hypoxic seedlings were characterized by the up-regulation of glutamate dehydrogenase 1 (GDH1) and mitochondrial alanine aminotransferase (mAlaAT), and down-regulation of glutamine synthetase 1b (GS1b), NADH-glutamate synthase (NADH-GOGAT), glutamate dehydrogenase 3 (GDH3), and isocitrate dehydrogenase (ICDH) gene expression. Hypoxic stress severely inhibited GS activity and stimulated NADH-GOGAT activity. GDH activity was lower in hypoxic seedlings than in the control, however, under either normoxia or hypoxia, the in vivo activity was directed towards glutamate deamination. (15)NH(4) labelling showed for the first time that the adaptive reaction of the plant to hypoxia consisted of a concerted modulation of nitrogen flux through the pathways of both alanine and glutamate synthesis. In hypoxic seedlings, newly synthesized (15)N-alanine increased and accumulated as the major amino acid, asparagine synthesis was inhibited, while (15)N-glutamate was synthesized at a similar rate to that in the control. A discrepancy between the up-regulation of GDH1 expression and the down-regulation of GDH activity by hypoxic stress highlighted for the first time the complex regulation of this enzyme by hypoxia. Higher rates of glycolysis and ethanol fermentation are known to cause the fast depletion of sugar stores and carbon stress. It is proposed that the expression of GDH1 was stimulated by hypoxia-induced carbon stress, while the enzyme protein might be involved during post-hypoxic stress contributing to the regeneration of 2-oxoglutarate via the GDH shunt.     

Genetic and physiological analysis of germination efficiency in maize in relation to nitrogen metabolism reveals the importance of cytosolic glutamine synthetase

We have developed an approach combining physiology and quantitative genetics to enhance our understanding of nitrogen (N) metabolism during kernel germination. The physiological study highlighted the central role of glutamine (Gln) synthetase (GS) and Gln synthesis during this developmental process because a concomitant increase of both the enzyme activity and the amino acid content was observed. This result suggests that Gln is acting either as a sink for ammonium released during both storage protein degradation and amino acid deamination or as a source for amino acid de novo synthesis by transamination. In the two parental lines used for the quantitative genetics approach, we found that the increase in Gln occurred earlier in Io compared with F(2), a result consistent with its faster germinating capacity. The genetic study was carried out on 140 F6 recombinant inbred lines derived from the cross between F(2) and Io. Quantitative trait locus mapping identified three quantitative trait loci (QTLs) related to germination trait (T50, time at which 50% of the kernels germinated) that explain 18.2% of the phenotypic variance; three QTLs related to a trait linked to germination performance, kernel size/weight (thousand kernels weight), that explain 17% of the phenotypic variance; two QTLs related to GS activity at early stages of germination that explain 17.7% of the phenotypic variance; and one QTL related to GS activity at late stages of germination that explains 7.3% of the phenotypic variance. Coincidences of QTL for germination efficiency and its components with genes encoding cytosolic GS (GS1) and the corresponding enzyme activity were detected, confirming the important role of the enzyme during the germination process. A triple colocalization on chromosome 4 between gln3 (a structural gene encoding GS1) and a QTL for GS activity and T50 was found; whereas on chromosome 5, a QTL for GS activity and thousand kernels weight colocalized with gln4, another structural gene encoding GS1. This observation suggests that for each gene, the corresponding enzyme activity is of major importance for germination efficiency either through the size of the grain or through its faster germinating capacity. Consistent with the possible nonoverlapping function of the two GS1 genes, we found that in the parental line Io, the expression of Gln3 was transiently enhanced during the first hours of germination, whereas that of gln4 was constitutive.