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Hepatic glutamine synthetase controls N⁵-methylglutamine in homeostasis and cancer.

ABSTRACT: Glutamine synthetase (GS) activity is conserved from prokaryotes to humans, where the ATP-dependent production of glutamine from glutamate and ammonia is essential for neurotransmission and ammonia detoxification. Here, we show that mammalian GS uses glutamate and methylamine to produce a methylated glutamine analog, N⁵-methylglutamine. Untargeted metabolomics revealed that liver-specific GS deletion and its pharmacological inhibition in mice suppress hepatic and circulating levels of N⁵-methylglutamine. This alternative activity of GS was confirmed in human recombinant enzyme and cells, where a pathogenic mutation in the active site (R324C) promoted the synthesis of N⁵-methylglutamine over glutamine. N⁵-methylglutamine is detected in the circulation, and its levels are sustained by the microbiome, as demonstrated by using germ-free mice. Finally, we show that urine levels of N⁵-methylglutamine correlate with tumor burden and GS expression in a β-catenin-driven model of liver cancer, highlighting the translational potential of this uncharacterized metabolite.

SUBMITTER: Villar VH

PROVIDER: S-EPMC9974483 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature

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Hepatic glutamine synthetase controls N<sup>5</sup>-methylglutamine in homeostasis and cancer.

Villar Victor H VH Allega Maria Francesca MF Deshmukh Ruhi R Ackermann Tobias T Nakasone Mark A MA Vande Voorde Johan J Drake Thomas M TM Oetjen Janina J Bloom Algernon A Nixon Colin C Müller Miryam M May Stephanie S Tan Ee Hong EH Vereecke Lars L Jans Maude M Blancke Gillian G Murphy Daniel J DJ Huang Danny T DT Lewis David Y DY Bird Thomas G TG Sansom Owen J OJ Blyth Karen K Sumpton David D Tardito Saverio S

Nature chemical biology 20221024 3

Glutamine synthetase (GS) activity is conserved from prokaryotes to humans, where the ATP-dependent production of glutamine from glutamate and ammonia is essential for neurotransmission and ammonia detoxification. Here, we show that mammalian GS uses glutamate and methylamine to produce a methylated glutamine analog, N<sup>5</sup>-methylglutamine. Untargeted metabolomics revealed that liver-specific GS deletion and its pharmacological inhibition in mice suppress hepatic and circulating levels of ...[more]

PMID: 36280791

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Project description:BackgroundHepatic fibrosis is a common pathologic stage in chronic liver disease development, which might ultimately lead to liver cirrhosis. Accumulating evidence suggests that adipose-derived stromal cells (ADSCs)-based therapies show excellent therapeutic potential in liver injury disease owing to its superior properties, including tissue repair ability and immunomodulation effect. However, cell-based therapy still limits to several problems, such as engraftment efficiency and immunoreaction, which impede the ADSCs-based therapeutics development. So, ADSCs-derived extracellular vesicles (EVs), especially for exosomes (ADSC-EXO), emerge as a promise cell-free therapeutics to ameliorate liver fibrosis. The effect and underlying mechanisms of ADSC-EXO in liver fibrosis remains blurred.MethodsHepatic fibrosis murine model was established by intraperitoneal sequential injecting the diethylnitrosamine (DEN) for two weeks and then carbon tetrachloride (CCl4) for six weeks. Subsequently, hepatic fibrosis mice were administrated with ADSC-EXO (10 μg/g) or PBS through tail vein infusion for three times in two weeks. To evaluate the anti-fibrotic capacity of ADSC-EXO, we detected liver morphology by histopathological examination, ECM deposition by serology test and Sirius Red staining, profibrogenic markers by qRT-PCR assay. LX-2 cells treated with TGF-β (10 ng/ml) for 12 h were conducted for evaluating ADSC-EXO effect on activated hepatic stellate cells (HSCs). RNA-seq was performed for further analysis of the underlying regulatory mechanisms of ADSC-EXO in liver fibrosis.ResultsIn this study, we obtained isolated ADSCs, collected and separated ADSCs-derived exosomes. We found that ADSC-EXO treatment could efficiently ameliorate DEN/CCl4-induced hepatic fibrosis by improving mice liver function and lessening hepatic ECM deposition. Moreover, ADSC-EXO intervention could reverse profibrogenic phenotypes both in vivo and in vitro, including HSCs activation depressed and profibrogenic markers inhibition. Additionally, RNA-seq analysis further determined that decreased glutamine synthetase (Glul) of perivenous hepatocytes in hepatic fibrosis mice could be dramatically up-regulated by ADSC-EXO treatment; meanwhile, glutamine and ammonia metabolism-associated key enzyme OAT was up-regulated and GLS2 was down-regulated by ADSC-EXO treatment in mice liver. In addition, glutamine synthetase inhibitor would erase ADSC-EXO therapeutic effect on hepatic fibrosis.ConclusionsThese findings demonstrated that ADSC-derived exosomes could efficiently alleviate hepatic fibrosis by suppressing HSCs activation and remodeling glutamine and ammonia metabolism mediated by hepatocellular glutamine synthetase, which might be a novel and promising anti-fibrotic therapeutics for hepatic fibrosis disease.

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Hepatic glutamine synthetase controls N⁵-methylglutamine in homeostasis and cancer.

Publications

Hepatic glutamine synthetase controls N<sup>5</sup>-methylglutamine in homeostasis and cancer.

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Hepatic glutamine synthetase controls N5-methylglutamine in homeostasis and cancer.

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Hepatic glutamine synthetase controls N&lt;sup&gt;5&lt;/sup&gt;-methylglutamine in homeostasis and cancer.

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Hepatic glutamine synthetase controls N⁵-methylglutamine in homeostasis and cancer.

Hepatic glutamine synthetase controls N<sup>5</sup>-methylglutamine in homeostasis and cancer.