Gamma-Glutamyl transpeptidase in rat mammary tissue. Activity during lactogenesis and regulation by prolactin.
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ABSTRACT:
SUBMITTER: Pocius PA
PROVIDER: S-EPMC1161904 | biostudies-other | 1980 May
REPOSITORIES: biostudies-other
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Project description:γ-Glutamyl transpeptidase (GGT) is an enzyme located on the surface of cellular membranes and involved in GSH metabolism and maintenance of redox homeostasis. High GGT expression on tumor cells is associated with increased cell proliferation and resistance against chemotherapy. GGT inhibitors evaluated so far in clinical trials are too toxic for human use. In this study, using enzyme kinetics analyses, we demonstrate that ovothiols, 5(Nπ)-methyl thiohistidines of marine origin, act as noncompetitive inhibitors of GGT, with an apparent Ki of 21 μm, when we fixed the concentrations of the donor substrate. We found that these compounds are more potent than the known GGT inhibitor 6-diazo-5-oxo-l-norleucine and are not toxic toward human embryonic cells. In particular, cellular process-specific fluorescence-based assays revealed that ovothiols induce a mixed cell-death phenotype of apoptosis and autophagy in GGT-overexpressing cell lines, including human liver cancer and chronic B leukemic cells. The findings of our study provide the basis for further development of 5-thiohistidines as therapeutics for GGT-positive tumors and highlight that GGT inhibition is involved in autophagy.
| S-EPMC6779435 | biostudies-literature
Project description:Background and aimCoronavirus disease 2019 (COVID-19) is a life-threatening disease that predominantly causes respiratory failure. The impact of COVID-19 on other organs remains elusive. Herein, we aimed to investigate the effects of COVID-19 on the hepatobiliary system.MethodsIn the current study, we obtained the clinical records and laboratory results from 66 laboratory-confirmed patients with COVID-19 at the Wuhan Tongji Hospital between 10 February 2020 and 28 February 2020. The detailed clinical features and laboratory findings were collected for analysis. Bioinformatics analysis was conducted to evaluate the correlation between gamma-glutamyl transferase (GGT) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry receptor angiotensin-converting enzyme 2 (ACE2).ResultsIn this cohort, 30 (51.7%) patients had abnormal liver function on admission, which was associated with disease severity and enriched in the male and diabetic patients. The elevated levels of direct bilirubin (P = 0.029) and GGT (P = 0.004) were common in patients with severe pneumonia when compared with those with mild pneumonia. In addition, elevated levels of GGT (P = 0.003) and aspartate aminotransferase (AST) (P = 0.007) were positively associated with longer hospital stay. The expression of ACE2 was closely associated with GGT in various human tissues because they shared the common transcriptional regulator hepatic nuclear factor-1β (HNF1B).ConclusionsIncreased GGT levels were common in severe cases and elevated GGT levels were positively associated with prolonged hospital stay and disease severity. Due to the consistent expression with ACE2, GGT is a potent biomarker indicating the susceptibility of SARS-CoV-2 infection.
| S-EPMC8464026 | biostudies-literature
Project description:Myotonic dystrophy-related Cdc42-binding kinase alpha (MRCKα) is an integral component of signaling pathways controlling vital cellular processes, including cytoskeletal reorganization, cell proliferation and cell survival. In this study, we investigated the physiological role of MRCKα in milk protein and fat production in dairy cows, which requires a dynamic and strict organization of the cytoskeletal network in bovine mammary epithelial cells (BMEC). Within a selection of 9 Holstein cows, we found that both mRNA and protein expression of MRCKα in the mammary gland were upregulated during lactation and correlated positively (r > 0.89) with the mRNA and protein levels of β-casein. Similar positive correlations (r > 0.79) were found in a primary culture of BMEC stimulated with prolactin for 24 h. In these cells, silencing of MRCKα decreased basal β-casein, sterol-regulatory element binding protein (SREBP)-1 and cyclin D1 protein level, phosphorylation of mTOR, triglyceride secretion, cell number and viability-while overexpression of MRCKα displayed the reversed effect. Notably, silencing of MRCKα completely prevented the stimulatory action of prolactin on the same parameters. These data demonstrate that MRCKα is a critical mediator of prolactin-induced lactogenesis via stimulation of the mTOR/SREBP1/cyclin D1 signaling pathway.
| S-EPMC9304597 | biostudies-literature
Project description:The genetic hallmark of Ewing's sarcoma family of tumours (ET) is the presence of the translocation t(11;22)(q24;q12), which creates the ET fusion gene, leading to cellular transformation. Five human gamma-glutamyl transpeptidase (gamma-GT) genes are located near the chromosomal translocation in ET. gamma-GT is a major enzyme involved in glutathione homoeostasis. Five human cell lines representative of primary or metastatic tumours were investigated to study whether gamma-GT alterations could occur at the chromosomal breaks and rearrangements in ET. As shown by enzymic assays and FACS analyses, all ET cell lines consistently expressed a functional gamma-GT which however did not discriminate steps of ET progression. As shown previously [Sancéau, Hiscott, Delattre and Wietzerbin (2000) Oncogene 19, 3372-3383], ET cells respond to the antiproliferative effects of interferons (IFNs) type I (alpha and beta) and to a much less degree to IFN type II (gamma). IFN-alpha and -beta arrested cells in the S-phase of the cell cycle. We found an enhancement of gamma-GT mRNA species with IFN-alpha and -beta by reverse transcriptase-PCR analyses. This is reflected by up-regulation of gamma-GT protein, which coincides with the increase in gamma-GT-specific enzymic activity. Similarly, IFNs up-regulate the levels of gamma-GT in another IFN-responsive B cell line. Whether this up-regulation of gamma-GT by IFNs is of physiological relevance to cell behaviour remains to be studied.
| S-EPMC1222621 | biostudies-other
Project description:Monoclonal antibodies have been raised against rat kidney gamma-glutamyl transpeptidase (GGT). All five antibodies immunoprecipitate enzyme activity from solubilized kidney brush-border membranes, but not from hepatocellular carcinoma membranes. Three of the antibodies react immunohistochemically with brush-border membranes in sections of adult rat kidney, but none of the antibodies cross-react with sections of guinea-pig, mouse or marmoset kidney or with untreated or carcinogen-treated rat liver. The antibodies do not recognize GGT in foetal-rat kidney and react poorly with kidney from 2-year-old rat. They do react with tubules trapped within mesenchymal kidney tumours induced by dimethylnitrosamine, but epithelial tumours, which are GGT-positive (although much less so than normal kidney), are not immunoreactive.
| S-EPMC1147221 | biostudies-other
Project description:To investigate the role of γ-glutamyltranspeptidase in severe asthma, we established an OVA/ LPS-induced asthma model. Inflammation analysis and second-generation sequencing revealed the mechanism by which inhibition of γ-glutamyltranspeptidase inhibited LPS-induced asthma exacerbation in mice.
2023-07-05 | GSE217834 | GEO
Project description:Expression of gamma-glutamyl transpeptidase (GGT) in tumors contributes to resistance to radiation and chemotherapy. GGT is inhibited by glutamine analogues that compete with the substrate for the gamma-glutamyl binding site. However, the glutamine analogues that have been evaluated in clinical trials are too toxic for use in humans. We have used high throughput screening to evaluate small molecules for their ability to inhibit GGT and have identified a novel class of inhibitors that are not glutamine analogues. These compounds are uncompetitive inhibitors, binding the gamma-glutamyl enzyme complex. OU749, the lead compound, has an intrinsic K(i) of 17.6 microm. It is a competitive inhibitor of the acceptor glycyl-glycine, which indicates that OU749 occupies the acceptor site while binding to the gamma-glutamyl substrate complex. OU749 is more than 150-fold less toxic than the GGT inhibitor acivicin toward dividing cells. Inhibition of GGT by OU749 is species-specific, inhibiting GGT isolated from human kidney with 7-10-fold greater potency than GGT isolated from rat or mouse kidney. OU749 does not inhibit GGT from pig cells. Human GGT expressed in mouse fibroblasts is inhibited by OU749 similarly to GGT from human cells, which indicates that the species specificity is determined by differences in the primary structure of the protein rather than species-specific, post-translational modifications. These studies have identified a novel class of inhibitors of GGT, providing the basis for further development of a new group of therapeutics that inhibit GGT by a mechanism distinct from the toxic glutamine analogues.
| S-EPMC2666554 | biostudies-literature
Project description:gamma-Glutamyl transpeptidase (EC 2.3.2.2, gamma GT) is a membrane-bound ectoenzyme that plays an important role in the metabolism of glutathione. It is composed of two subunits, both of which are encoded by a common mRNA. We examined the expression of gamma GT in human lung tissue by Northern blot analysis and screening a cDNA library made from human lung poly(A)+ RNA. Our results show that there are two gamma GT mRNA populations in human lung tissue. We define these as group I (2.4 kb) and group II (approximately 1.2 kb) transcripts. In the present communication, we characterize the unique lung transcript. Sequence analysis of representative clones shows that group I transcripts are virtually identical to those previously isolated from liver and placenta but possess a unique 5' untranslated region. In marked contrast, group II transcripts appear to be human-lung-specific. Group II transcripts appear on Northern blots probed with full-length or 3'-biased gamma GT cDNA. Sequence analysis of group II clones shows them to be homologous with group I clones in the region that encodes the reading frame for the light chain; however, they possess a series of unique 5' untranslated regions, which suggests that they arise from lung-specific message processing. Additionally, approximately 50% of the isolated group II clones contain 34 nt substitutions compared with the "wild-type" gamma GT transcripts. These data indicate that human lung expresses unique gamma GT transcripts of unknown function as well as the classical form. The abundant group II transcripts may encode part of a heterodimer related to gamma GT or represent processed lung-specific pseudogenes.
| S-EPMC47161 | biostudies-other
Project description:Backgroundγ-glutamyltranspeptidase (GGT) is a bi-substrate enzyme conserved in all three domains of life. It catalyzes the cleavage and transfer of γ-glutamyl moiety of glutathione to either water (hydrolysis) or substrates like peptides (transpeptidation). GGTs exhibit great variability in their enzyme kinetics although the mechanism of catalysis is conserved. Recently, GGT has been shown to be a virulence factor in microbes like Helicobacter pylori and Bacillus anthracis. In mammalian cells also, GGT inhibition prior to chemotherapy has been shown to sensitize tumors to the therapy. Therefore, lately both bacterial and eukaryotic GGTs have emerged as potential drug targets, but the efforts directed towards finding suitable inhibitors have not yielded any significant results yet. We propose that delineating the residues responsible for the functional diversity associated with these proteins could help in design of species/clade specific inhibitors.ResultsIn the present study, we have carried out phylogenetic analysis on a set of 47 GGT-like proteins to address the functional diversity. These proteins segregate into various subfamilies, forming separate clades on the tree. Sequence conservation and motif prediction studies show that even though most of the highly conserved residues have been characterized biochemically in previous studies, a significant number of novel putative sites and motifs are discovered that vary in a clade specific manner. Many of the putative sites predicted during the functional divergence type I and type II analysis, lie close to the known catalytic residues and line the walls of the substrate binding cavity, reinforcing their role in modulating the substrate specificity, catalytic rates and stability of this protein.ConclusionThe study offers interesting insights into the evolution of GGT-like proteins in pathogenic vs. non-pathogenic bacteria, archaea and eukaryotes. Our analysis delineates residues that are highly specific to each GGT subfamily. We propose that these sites not only explain the differences in stability and catalytic variability of various GGTs but can also aid in design of specific inhibitors against particular GGTs. Thus, apart from the commonly used in-silico inhibitor screening approaches, evolutionary analysis identifying the functional divergence hotspots in GGT proteins could augment the structure based drug design approaches.
| S-EPMC4568574 | biostudies-literature
Project description:The antioxidant properties of glutathione (GSH) and their relevance to oxidative stress induced pathological states such as Alzheimer's disease is well-established. The utility of GSH itself as a pharmacotherapeutic agent for such disorders is limited because of the former's lability to breakdown through amide cleavage by the ubiquitous enzyme γ-glutamyl transpeptidase (γ-GT). In the present study, a GSH analogue, Ψ-GSH, where the γ-glutamylcysteine amide linkage is replaced with a ureide linkage, was synthesized. Ψ-GSH was found to be stable toward γ-GT mediated breakdown. Ψ-GSH fulfilled four cardinal properties of GSH, namely, traversing across the blood brain barrier (BBB) via the GSH active uptake machinery, replacing GSH in the glyoxalase-I mediated detoxification of methylglyoxal, protecting cells against chemical oxidative insult, and finally lowering the cytotoxicity of amyloid-β peptide. These results validate Ψ-GSH as a viable metabolically stable replacement for GSH and establish it as a potential preclinical candidate for treatment of oxidative stress mediated pathology.
| S-EPMC3369799 | biostudies-literature