Evidence supporting distinct functions of three cytosolic glutamine synthetases and two NADH-glutamate synthases in rice

Abstract

The functions of the three isoenzymes of cytosolic glutamine synthetase (GS1;1, GS1;2, and GS1;3) and two NADH-glutamate synthases (NADH-GOGAT1 and NADH-GOGAT2) in rice (Oryza sativa L.) were characterized using a reverse genetics approach and spatial expression of the corresponding genes. OsGS1;2 and OsNADH-GOGAT1 were mainly expressed in surface cells of rice roots in an NH4 (+)-dependent manner. Disruption of either gene by the insertion of endogenous retrotransposon Tos17 caused reduction in active tiller number and hence panicle number at harvest. Re-introduction of OsGS1;2 cDNA under the control of its own promoter into the knockout mutants successfully restored panicle number to wild-type levels. These results indicate that GS1;2 and NADH-GOGAT1 are important in the primary assimilation of NH4 (+) taken up by rice roots. OsGS1;1 and OsNADH-GOGAT2 were mainly expressed in vascular tissues of mature leaf blades. OsGS1;1 mutants showed severe reduction in growth rate and grain filling, whereas OsNADH-GOGAT2 mutants had marked reduction in spikelet number per panicle. Complementation of phenotypes seen in the OsGS1;1 mutant was successfully observed when OsGS1;1 was re-introduced. Thus, these two enzymes could be important in remobilization of nitrogen during natural senescence. Metabolite profiling data showed a crucial role of GS1;1 in coordinating metabolic balance in rice. Expression of OsGS1:3 was spikelet-specific, indicating that it is probably important in grain ripening and/or germination. Thus, these isoenzymes seem to possess distinct and non-overlapping functions and none was able to compensate for the individual function of another.

Keywords: Glutamate synthase; glutamine synthetase; metabolic balance; nitrogen utilization; physiological function; rice..