NADH-GOGAT is a minor form of GOGAT species and little has been known concerning its cellular and subcellular localization, gene structure and expression, or function in higher plants. In 1992, a specific antibody for NADH-GOGAT from rice cell cultures was prepared as the first instance to crossreact NADH-GOGAT protein in rice plants.19 Antibody for NADH-GOGAT in alfalfa root nodules had been obtained prior to the rice, but the antibody hardly recognized NADH-GOGAT in plant organs other than nodules.20 Using the antibody for rice NADH-GOGAT, immunochemical and immunocyto-logical experiments have been performed. A high abundance of NADH-GOGAT protein was detected in the non-green, developing leaf blade14 and in the developing grains.21 Immunocytological results showed that the NADH-GOGAT protein was present in vascular parenchyma cells and mestome sheath cells of vascular bundles of the developing leaf blade, as well as in vascular parenchyma cells, nucellar projection, and nucellar epidermis of dorsal vascular bundles of the developing young grains.22 On the other hand, Fd- GOGAT protein is mainly present in mesophyll cells of leaf blades and in the chloroplast-containing cross cells of the pericarp of the grains.22 The spatial localization of these GOGAT proteins indicates distinct and non-overlapping roles, as in the case of GS isoenzymes. The cells containing NADH- GOGAT protein in leaf vascular bundles and the dorsal vascular bundles of grains are considered by the anatomical studies to be active in the solute transport from phloem and xylem.17 In addition, the content of NADH- GOGAT protein, as well as its activity, in the apical spikelets increased several-fold in the first 2 weeks after flowering.21 Thus, it is likely that NADH-GOGAT is involved in the re-utilization of glutamine transported through the vascular system, and the synthesized glutamate is further utilized for biosynthetic reactions in these young organs.
We have recently determined the gene structure for NADH-GOGAT in rice plants.23 To obtain direct evidence to support the possible role of NADH-GOGAT in nitrogen recycling, a cauliflower mosaic virus 35S promoter was fused with a fragment of NADH-GOGAT cDNA in the antisense orientation, and the chimeric construct was introduced into rice calli by the methods of Agrobacterium-mediated transformation.24 The experiments to look at the effects of expression of the antisense RNA in these transformants are now in progress. We have just obtained results which show that the weight of 1,000 grains from some of the transformants at the T0 generation is significantly reduced when compared to the grain weight of the control transformants introduced to vector without the NADH-GOGAT cDNA fragment (Fig. 17.2).
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