Disease relevance of GCLM

• The modifier subunit of glutamate cysteine ligase (GCLM) is a molecular target for amelioration of cisplatin resistance in lung cancer [1].
• METHODS AND RESULTS: We searched for the common variants in the 5'-flanking region of the GCL modifier subunit (GCLM) gene in patients with myocardial infarction (MI) [2].
• Recombinant hGLCLR andhGLCLC were produced in cells infected with recombinant baculoviruses, and homogeneous hGLCL subunits and holoenzyme were purified from cell lysates with a Ni-NTA resin [3].
• Squamous cell carcinoma xenografts significantly showed higher expression of GCLM gene than adenocarcinoma xenografts (p=0.023, t-test), while there was no significant difference in GCLC gene expression levels between each histopathological xenograft [4].
• Therefore we conducted a series of studies using rats and a human hepatoma (HepG2/C3A) cell line to assess the role of GCLM and holoenzyme formation in the regulation of GCL activity in response to sulphur amino acid intake or availability [5].

High impact information on GCLM

• GCLM gene expression is decreased in patients' fibroblasts [6].
• Inhibition of gamma-glutamylcysteine synthetase (gammaGCS) by buthionine sulfoximine augmented the increase in islet peroxide and decrease in insulin mRNA levels, content, and secretion in islets and HIT-T15 cells induced by ribose [7].
• A mutant of Escherichia coli, JTG10, deficient in gamma-glutamylcysteine synthetase (gamma-ECS; EC 6.3.2.2) is unable to synthesize glutathione (GSH) and is sensitive to 8-hydroxyquinoline [8].
• This phenotype was exploited for the isolation of Arabidopsis thaliana gamma-ECS cDNAs by expression cloning, and clones were selected through functional complementation by growth on 8-hydroxyquinoline [8].
• The activity of recombinant Arabidopsis gamma-ECS was inhibited by buthionine sulfoximine and GSH, indicating that, while differences in the primary and secondary structure of gamma-ECS from different sources exist, the enzymes may have similar active site structures [8].

Chemical compound and disease context of GCLM

• BACKGROUND: The minor -588T allele of polymorphism -588C/T of a modifier subunit gene in glutamate-cysteine ligase (GCLM), a rate-limiting enzyme for glutathione (GSH) synthesis, was associated with lower plasma GSH levels and was a risk factor for myocardial infarction [9].
• Structure and refinement of the physical mapping of the gamma- glutamylcysteine ligase regulatory subunit (GLCLR) gene to chromosome 1p22.1 within the critically deleted region of human malignant mesothelioma [10].
• Transient induction of the MRP/GS-X pump and gamma-glutamylcysteine synthetase by 1-(4-amino-2-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3- nitrosourea in human glioma cells [11].
• In a series of ovarian carcinoma cell lines selected in vitro for resistance to cisplatin by continuous exposure to increasing drug concentrations, the level of resistance is proportional to the expression of gamma-glutamylcysteine synthetase (gamma-GCS) [12].
• Here we investigated whether L-buthionine-(S,R)-sulphoximine (BSO), an inhibitor of gamma-glutamylcysteine synthetase, alters endothelial and vascular smooth muscle injury in response to peroxynitrite in vitro and during endotoxic shock in vivo [13].

Biological context of GCLM

• GCL is the rate-limiting step and represents a heterodimeric enzyme comprised of a catalytic subunit (GCLC) and a ("regulatory"), or modifier, subunit (GCLM) [14].
• Curcumin also rapidly stimulated PKC phosphorylation and Ro-31-8220, a pan-PKC inhibitor, decreased curcumin-induced GCLM and HO-1 mRNA expression and ARE binding [15].
• To understand better the structure-function relationship of GLCL subunits and holoenzyme, human GLCLR and GLCLC genes were inserted into the baculovirus genome [3].
• Here we specify the chromosomal sublocalization of the human gene (GLCLR) that encodes the regulatory subunit of E.C. 6.3.2.2 to chromosome 1p21 [16].
• We recently assigned the human gene (GLCLC) that encodes the catalytic subunit of gamma-glutamylcysteine synthetase (glutamate-cysteine ligase, E.C. 6.3.2.2) to human chromosome 6p12 [16].

Anatomical context of GCLM

• CONCLUSIONS: The -588T polymorphism of the GCLM gene causes a decrease in endothelial NO bioactivity, leading to impairment of endothelium-dependent vasomotor function in large and resistance coronary arteries [9].
• Regulation of gamma-glutamylcysteine synthetase regulatory subunit (GLCLR) gene expression: identification of the major transcriptional start site in HT29 cells [17].
• Bleomycin upregulates expression of gamma-glutamylcysteine synthetase in pulmonary artery endothelial cells [18].
• Glutathione depletion achieved by continuous exposure of mitogen-activated human T lymphocytes to L-buthionine-(S,R)-sulfoximine, a specific inhibitor of gamma-glutamylcysteine synthetase, leads to a marked inhibition of the proliferative response [19].
• Selenite addition increased the activity of gamma-glutamylcysteine synthetase in canine mammary tumor cell line 11 and non-neoplastic canine mammary cells, but not in canine mammary tumor cell line 13 cells [20].

Associations of GCLM with chemical compounds

• GCLC owns the catalytic activity, whereas GCLM enhances the enzyme activity by lowering the K(m) for glutamate and increasing the K(i) to GSH inhibition [14].
• Rottlerin (a PKC delta inhibitor) and PKC delta antisense oligonucleotides significantly inhibited curcumin-induced GCLM and HO-1 mRNA expression and ARE binding [15].
• Glutamate-L-cysteine ligase (GLCL [EC 6.3.2.2], also referred to as gamma-glutamylcysteine synthetase) catalyzes the rate-limiting reaction in the synthesis of the important cellular antioxidant glutathione [21].
• The transcriptional activity of the GLCLR gene was increased by approximately 2.5-fold in SNP-treated cells [17].
• Pyrrolidine dithiocarbamate (PDTC) induction of the human glutamate cysteine ligase modulatory (GCLM) gene is dependent on activation of the mitogen-activated protein kinases (MAPKs) extracellular regulated kinase (Erk) and p38, and is not affected by protein kinase C (PKC) or PI3K inhibitors [22].

Regulatory relationships of GCLM

• CYP2E1 overexpression in HepG2 cells induces glutathione synthesis by transcriptional activation of gamma-glutamylcysteine synthetase [23].

Other interactions of GCLM

• GCL is a heterodimeric protein with a catalytic (or heavy, GCLC) subunit and a modulatory (or light, GCLM) subunit [24].
• GCLM and TXNRD1 exhibited 13- and 9-fold increases, respectively at 24 h [25].
• Delayed mechanism for induction of gamma-glutamylcysteine synthetase heavy subunit mRNA stability by oxidative stress involving p38 mitogen-activated protein kinase signaling [26].
• Growth inhibition of P. falciparum 3D7 by D,L-buthionine-( S, R )sulphoximine (BSO), an inhibitor of gamma-glutamylcysteine synthetase (gamma-GCS), and by Methylene Blue (MB), an inhibitor of gluta thione reductase (GR), was significantly more pronounced than inhibition of P. falciparum Dd2 growth by these drugs [27].
• Using genetic and chemical tools, it was demonstrated that thioredoxin reductase, the first step of the thioredoxin redox cycle, and gamma-glutamylcysteine synthetase (gamma-GCS), the rate-limiting step of glutathione synthesis, are essential for parasite survival [28].

Analytical, diagnostic and therapeutic context of GCLM

• There was a significant association between the expression of GCLM and GCLC mRNA in each xenograft (Fisher's test, p<0.045) [4].
• The gene of the key GSH-synthesizing enzyme, glutamate cysteine ligase modifier (GCLM) subunit, is strongly associated with schizophrenia in two case-control studies and in one family study [6].
• In situ hybridization and immunolocalization of leaf sections showed that gamma-ECS and GSH-S transcripts and proteins were found in both the bundle sheath (BS) and the mesophyll cells under optimal conditions [29].
• The mRNA and protein for the rate-limiting enzyme for GSH synthesis, gamma-glutamylcysteine synthetase (GCS), were also determined in alphaA- and mock-transfected cells by Northern blot analysis and Western blot analysis of heavy (GCS-HS) and light (GCS-LS) subunits [30].
• Sequence analysis of the promoter region demonstrated the presence of putative enhancer elements including AP-1 sites and an antioxidant-responsive element, which might be involved in the observed induction of the GLCLR promoter [31].

References