Resolving the Role of Plant Glutamate Dehydrogenase. I. in vivo Real Time Nuclear Magnetic Resonance Spectroscopy Experiments

In higher plants the glutamate dehydrogenase (GDH) enzyme catalyzes the reversible amination of 2-oxoglutarate to form glutamate, using ammonium as a substrate. For a better understanding of the physiological function of GDH either in ammonium assimilation or in the supply of 2-oxoglutarate, we used transgenic tobacco (Nicotiana tabacum L.) plants overexpressing the two genes encoding the enzyme. An in vivo real time 15N-nuclear magnetic resonance (NMR) spectroscopy approach allowed the demonstration that, when the two GDH genes were overexpressed individually or simultaneously, the transgenic plant leaves did not synthesize glutamate in the presence of ammonium when glutamine synthetase (GS) was inhibited. In contrast we confirmed that the primary function of GDH is to deaminate Glu. When the two GDH unlabeled substrates ammonium and Glu were provided simultaneously with either [15N]Glu or 15NH4+ respectively, we found that the ammonium released from the deamination of Glu was reassimilated by the enzyme GS, suggesting the occurrence of a futile cycle recycling both ammonium and Glu. Taken together, these results strongly suggest that the GDH enzyme, in conjunction with NADH-GOGAT, contributes to the control of leaf Glu homeostasis, an amino acid that plays a central signaling and metabolic role at the interface of the carbon and nitrogen assimilatory pathways. Thus, in vivo NMR spectroscopy appears to be an attractive technique to follow the flux of metabolites in both normal and genetically modified plants

Biosynthesis and secretion of cryptogein, a protein elicitor secreted by Phytophthora cryptogea

International audienc

The Key Role of Glutamate Dehydrogenase 2 (GDH2) in the Control of Kernel Production in Maize ( Zea mays L.)

The agronomic potential of glutamate dehydrogenase 2 (GDH2) in maize kernel production was investigated by examining the impact of a mutation on the corresponding gene. Mu-insertion homozygous and heterozygous mutant lines lacking GDH2 activity were isolated and characterized at the biochemical, physiological and agronomic levels. In comparison to the wild type and to the homozygous ghd2 mutants, the heterozygous gdh2 mutant plants were characterized by a decrease in the root amino acid content, whereas in the leaves an increase of a number of phenolic compounds was observed. On average, a 30 to 40% increase in kernel yield was obtained only in the heterozygous gdh2 mutant lines when plants were grown in the field over two years. The importance of GDH2 in the control of plant productivity is discussed in relation to the physiological impact of the mutation on amino acid content, with primary carbon metabolism mostly occurring in the roots and secondary metabolism occurring in the leaves

Use of H-1-NMR metabolomics to precise the function of the third glutamate dehydrogenase gene in Arabidopsis thaliana

It is now well established that the GS/GOGAT cycle is the major route for ammonium assimilation in higher plants. However, it has often been argued that other enzymes, such as glutamate dehydrogenase, have the capacity to assimilate ammonium, leading to the hypothesis that alternative ammonium assimilatory pathways could operate under particular physiological conditions. The GDH enzyme is encoded by two distinct genes, GDH1 and GDH2. A third gene, GDH3, potentially encoding GDH has recently been identified by in silico studies performed on Arabidopsis thaliana. In order to precise its function, the metabolic profile of gdh3 knock out mutants were compared to wild type plants using the 1H-NMR technique. 1H-NMR spectra coupled with principal component analysis and partial least square-discriminant analysis were applied to identify changes of the metabolic profiles. These experiments were performed on roots, leaves and stems. In the gdh3 mutant, metabolic variations were observed for carboxylic acids, amino acids and carbohydrates content

Glutamate Dehydrogenase of Tobacco Is Mainly Induced in the Cytosol of Phloem Companion Cells When Ammonia Is Provided Either Externally or Released during Photorespiration

Glutamate (Glu) dehydrogenase (GDH) catalyses the reversible amination of 2-oxoglutarate for the synthesis of Glu using ammonium as a substrate. This enzyme preferentially occurs in the mitochondria of companion cells of a number of plant species grown on nitrate as the sole nitrogen source. For a better understanding of the controversial role of GDH either in ammonium assimilation or in the supply of 2-oxoglutarate (F. Dubois, T. Tercé-Laforgue, M.B. Gonzalez-Moro, M.B. Estavillo, R. Sangwan, A. Gallais, B. Hirel [2003] Plant Physiol Biochem 41: 565–576), we studied the localization of GDH in untransformed tobacco (Nicotiana tabacum) plants grown either on low nitrate or on ammonium and in ferredoxin-dependent Glu synthase antisense plants. Production of GDH and its activity were strongly induced when plants were grown on ammonium as the sole nitrogen source. The induction mainly occurred in highly vascularized organs such as stems and midribs and was likely to be due to accumulation of phloem-translocated ammonium in the sap. GDH induction occurred when ammonia was applied externally to untransformed control plants or resulted from photorespiratory activity in transgenic plants down-regulated for ferredoxin-dependent Glu synthase. GDH was increased in the mitochondria and appeared in the cytosol of companion cells. Taken together, our results suggest that the enzyme plays a dual role in companion cells, either in the mitochondria when mineral nitrogen availability is low or in the cytosol when ammonium concentration increases above a certain threshold

Glutamate dehydrogenase in plants: is there a new story for an old enzyme

DOI: 10.1016/S0981-9428(03)00075-5International audienc

Control of the Synthesis and Subcellular Targeting of the Two GDH Genes Products in Leaves and Stems of Nicotiana plumbaginifolia and Arabidopsis thaliana

doi:10.1093/pcp/pcj008, available online at www.pcp.oupjournals.or