Abstract
Glutamine synthetase (E.C. 6.3.1.2) is a key enzyme of plant nitrogen metabolism that assimilates ammonia into glutamine. The Arabidopsis thaliana genome encodes one chloroplastic (GLN2) and five cytosolic (GLN1;1 ? GLN1;5) isoforms with different expression patterns, kinetic properties, regulation and functions. Physiological roles of different isoforms have been elucidated mainly by studying knockout mutants. However, the role of GLN1;5, which is expressed in dry seeds, remains unknown. To clarifty the function of GLN1;5, we studied a GLN1;5 knockout line (GLN1;5KO) homozygous for T-DNA insertion within the GLN1;5. GLN1;5 deficiency results in a phenotype with slightly delayed bolting and fewer siliques. The dry weight of GLN1;5KO seeds was 73.3% of wild-type (WT) seed weight, with seed length 90.9% of WT seeds. Finally, only 18.33% of the mutant seeds germinated in water within 10 days in comparison to 34.67% of WT seeds. KNO3 strongly stimulated germination of both GLN1;5KO and WT seeds, while germination in the presence of increasing NH4Cl concentrations potentiated the differences between the two genotypes. It can be concluded that GLN1;5 activity supports silique development and grain filling and that it has a role in ammonium reassimilation in the seed, as well as assimilation and/or detoxification of ammonium from the environment. [Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. ON173024 and Grant no. ON173015]
Highlights
Glutamine synthetase (GS or GLN, E.C. 6.3.1.2) is a central enzyme of plant nitrogen metabolism that catalyzes ATP-dependent assimilation of ammonia into glutamine, which serves as amido-donor and Ntransporting amino acid
Тo estimate the copy number of T-DNA insertions within the GLN1;5 knockout line (GLN1);5KO line, the DNA was digested with three restriction enzymes and Southern hybridization was performed using neomycin phosphotransferase II (NPTII) 362-bp fragment located near the right T-DNA border as a probe (Fig. 2)
The probe hybridized with 2-3 T-DNA insertions in the mutant genome depending on the used restriction enzyme
Summary
Glutamine synthetase (GS or GLN, E.C. 6.3.1.2) is a central enzyme of plant nitrogen metabolism that catalyzes ATP-dependent assimilation of ammonia into glutamine, which serves as amido-donor and Ntransporting amino acid. Arabidopsis GS1 genes have different spatiotemporal expression [3,4,5] and different kinetic properties [2] They are differentially regulated by both internal cues such as amino acids and carbon skeletons [6] and plant hormones [7], and by external signals, including ammonia [2], nitrates [8] and light [6,9]. For these reasons, specific and non-redundant functions have been suggested for Arabidopsis GS isoforms [1,4,10]
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