Cytosolic GLUTAMINE SYNTHETASE1;1 Modulates Metabolism and Chloroplast Development in Roots.

Nitrogen (N) is an essential macronutrient, and the final form of endogenous inorganic N is ammonium, which is assimilated by Gln synthetase (GS) into Gln. However, how the multiple isoforms of cytosolic GSs contribute to metabolic systems via the regulation of ammonium assimilation remains unclear. In this study, we compared the effects of two rice (Oryza sativa) cytosolic GSs, namely OsGS1;1 and OsGS1;2, on central metabolism in roots using reverse genetics, metabolomic and transcriptomic profiling, and network analyses. We observed (1) abnormal sugar and organic N accumulation and (2) significant up-regulation of genes associated with photosynthesis and chlorophyll biosynthesis in the roots of Osgs1;1 but not Osgs1;2 knockout mutants. Network analysis of the Osgs1;1 mutant suggested that metabolism of Gln was coordinated with the metabolic modules of sugar metabolism, tricarboxylic acid cycle, and carbon fixation. Transcript profiling of Osgs1;1 mutant roots revealed that expression of the rice sigma-factor (OsSIG) genes in the mutants were transiently upregulated. GOLDEN2-LIKE transcription factor-encoding genes, which are involved in chloroplast biogenesis in rice, could not compensate for the lack of OsSIGs in the Osgs1;1 mutant. Microscopic analysis revealed mature chloroplast development in Osgs1;1 roots but not in the roots of Osgs1;2, Osgs1;2-complemented lines, or the wild type. Thus, organic N assimilated by OsGS1;1 affects a broad range of metabolites and transcripts involved in maintaining metabolic homeostasis and plastid development in rice roots, whereas OsGS1;2 has a more specific role, affecting mainly amino acid homeostasis but not carbon metabolism.

Cytosolic glutamine synthetase1;2 is responsible for the primary assimilation of ammonium in rice roots.

Among three genes for cytosolic glutamine synthetase (OsGS1;1, OsGS1;2 and OsGS1;3) in rice (Oryza sativa L.) plants, the OsGS1;2 gene is known to be mainly expressed in surface cells of roots, but its function was not clearly understood. We characterized knock-out mutants caused by the insertion of an endogenous retrotransposon Tos17 into exon 2 of OsGS1;2. Homozygously inserted mutants showed severe reduction in active tiller number and hence panicle number at harvest. Other yield components, such as spikelet number per panicle, 1,000-spikelet weight and proportion of well ripened grains, were nearly identical between the mutants and wild-type plants. When the contents of free amino acids in roots were compared between the mutants and the wild type, there were marked reductions in contents of glutamine, glutamate, asparagine and aspartate, but a remarkable increase in free ammonium ions in the mutants. Concentrations of amino acids and ammonium ions in xylem sap behaved in a similar fashion. Re-introduction of OsGS1;2 cDNA under the control of its own promoter into the knock-out mutants successfully restored yield components to wild-type levels as well as ammonium concentration in xylem sap. The results indicate that GS1;2 is important in the primary assimilation of ammonium ions taken up by rice roots, with GS1;1 in the roots unable to compensate for GS1;2 functions.

Metabolomics data reveal a crucial role of cytosolic glutamine synthetase 1;1 in coordinating metabolic balance in rice.

Rice plants grown in paddy fields preferentially use ammonium as a source of inorganic nitrogen. Glutamine synthetase (GS) catalyses the conversion of ammonium to glutamine. Of the three genes encoding cytosolic GS in rice, OsGS1;1 is critical for normal growth and grain filling. However, the basis of its physiological function that may alter the rate of nitrogen assimilation and carbon metabolism within the context of metabolic networks remains unclear. To address this issue, we carried out quantitative comparative analyses between the metabolite profiles of a rice mutant lacking OsGS1;1 and its background wild type (WT). The mutant plants exhibited severe retardation of shoot growth in the presence of ammonium compared with the WT. Overaccumulation of free ammonium in the leaf sheath and roots of the mutant indicated the importance of OsGS1;1 for ammonium assimilation in both organs. The metabolite profiles of the mutant line revealed: (i) an imbalance in levels of sugars, amino acids and metabolites in the tricarboxylic acid cycle, and (ii) overaccumulation of secondary metabolites, particularly in the roots under a continuous supply of ammonium. Metabolite-to-metabolite correlation analysis revealed the presence of mutant-specific networks between tryptamine and other primary metabolites in the roots. These results demonstrated a crucial function of OsGS1;1 in coordinating the global metabolic network in rice plants grown using ammonium as the nitrogen source.