Glutamine synthetase expression in rat lung is regulated by protein stability

During physiological stress, the lung increases production of the amino acid glutamine (Gln) using the enzyme Gin synthetase (GS) to maintain Gin homeostasis. Glucocorticoid hormones are considered the principal mediators of GS expression during stress. However, whereas animal studies have shown that glucocorticoids increase lung GS mRNA levels 500-700%, GS activity levels rise only 20-45%. This discrepancy suggests that a posttranscriptional control mechanism(s) ultimately determines GS expression. We hypothesized that the level of GS protein in the lung is governed by the intracellular Gin concentration through a mechanism of protein destabilization, a feedback regulatory mechanism that has been observed in vitro. To test this hypothesis, Sprague-Dawley rats were treated with a Gin-free diet and the GS inhibitor methionine sulfoximine (MSO) to deplete tissue Gin levels and prevent this feedback regulation. Exposure to Gin-free chow and MSO (100 mg/kg body wt) for 6 days decreased plasma Gin levels 50% (P < 0.01) and decreased lung tissue Gin levels by 70% (P < 0.01). Although lung GS mRNA levels were not influenced by Gin depletion, there was a sevenfold (P < 0.01) increase in GS protein. A parenteral Gin infusion (200 mM, 1.5 ml/h) for the last 2 days of MSO treatment replenished lung Gin levels to 65% of control level and blunted the increase in GS protein levels by 33% (P < 0.05) compared with rats receiving an isomolar glycine solution. The acute effects of glucocorticoid and MSO administration on lung GS expression were also measured. Whereas dexamethasone (0.5 mg/kg) and MSO injections individually augmented lung GS protein levels twofold and fourfold (P < 0.05), respectively, the combination of dexamethasone and MSO produced a synergistic, 12-fold induction (P < 0.01) in lung GS protein over 8 h. The data suggest that, whereas glucocorticoids are potent mediators of GS transcriptional activity, protein stability greatly influences the ultimate expression of GS in the lung.