Glutamine (Gln), a major nitrogen source in plants, is considered a central intermediate that coordinates carbon-nitrogen assembly for plant growth and development. To maintain a sufficient Gln supply, plant cells employ glutamine synthetases (GS), including cytosolic GS1 and plastidic GS2 for Gln production. Previous work has shown that the <i>GS1</i> is responsive to various environmental stresses. This study demonstrated the involvement of <i>GS1</i>s in Gln homeostasis and the role of GS1 in abiotic stress tolerance in <i>Arabidopsis</i>. The <i>GS1</i> family is comprised of five isoforms in <i>Arabidopsis thaliana</i>. Gene expression profiling showed that <i>GLN1;1, GLN1;3</i> and <i>GLN1;4</i> had similar expression patterns and were upregulated by abiotic (salinity and cold) stresses, whereas <i>GLN1;2</i> exhibited constitutive expression and no <i>GLN1;5</i> transcript was detected under any of the conditions tested. Null T-DNA insertion mutants for the five <i>GS1</i> genes were obtained. Only the <i>gln1;1</i> mutant displayed enhanced sensitivity to a GS inhibitor, phosphinothricin, and to cold and salinity treatments, suggesting a nonredundant role for GLN1;1. Increased stress sensitivity in <i>gln1;1</i> was associated with accelerated accumulation of reactive oxygen species (ROS), particularly in chloroplasts. To better understand the role of cytosolic GS isoforms, we generated two different triple mutant combinations. Triple mutant <i>gln1;1/gln1;2/gln1;3</i> showed reduced growth at an early stage. The <i>gln1;1/gln1;3/gln1;4</i> mutant is pollen lethal, indicating an essential role of Gln in plant gametophyte development. Collectively, our results establish a link between cytosolic Gln production, ROS accumulation, plant stress tolerance and development.

Glutamine (Gln), a major nitrogen source in plants, is considered a central intermediate that coordinates carbon-nitrogen assembly for plant growth and development. To maintain a sufficient Gln supply, plant cells employ glutamine synthetases (GS), including cytosolic GS1 and plastidic GS2 for Gln production. Previous work has shown that the <i>GS1</i> is responsive to various environmental stresses. This study demonstrated the involvement of <i>GS1</i>s in Gln homeostasis and the role of GS1 in abiotic stress tolerance in <i>Arabidopsis</i>. The <i>GS1</i> family is comprised of five isoforms in <i>Arabidopsis thaliana</i>. Gene expression profiling showed that <i>GLN1;1, GLN1;3</i> and <i>GLN1;4</i> had similar expression patterns and were upregulated by abiotic (salinity and cold) stresses, whereas <i>GLN1;2</i> exhibited constitutive expression and no <i>GLN1;5</i> transcript was detected under any of the conditions tested. Null T-DNA insertion mutants for the five <i>GS1</i> genes were obtained. Only the <i>gln1;1</i> mutant displayed enhanced sensitivity to a GS inhibitor, phosphinothricin, and to cold and salinity treatments, suggesting a nonredundant role for GLN1;1. Increased stress sensitivity in <i>gln1;1</i> was associated with accelerated accumulation of reactive oxygen species (ROS), particularly in chloroplasts. To better understand the role of cytosolic GS isoforms, we generated two different triple mutant combinations. Triple mutant <i>gln1;1/gln1;2/gln1;3</i> showed reduced growth at an early stage. The <i>gln1;1/gln1;3/gln1;4</i> mutant is pollen lethal, indicating an essential role of Gln in plant gametophyte development. Collectively, our results establish a link between cytosolic Gln production, ROS accumulation, plant stress tolerance and development.