Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.5.1.18 (glutathione S-transferase)
22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The Bcl-2 family member Bcl-xL has often been correlated with apoptosis resistance. We have shown recently that in peripheral human T cells resistance to CD95-mediated apoptosis is characterized by a lack of caspase-8 recruitment to the CD95 death-inducing signaling complex (DISC) and by increased expression of Bcl-xL (Peter, M. E., Kischkel, F. C., Scheuerpflug, C. G., Medema, J. P., Debatin, K.-M., and Krammer, P. H. (1997) Eur. J. Immunol. 27, 1207-1212). This raises the possibility that Bcl-xL directly prevents caspase-8 activation by the DISC. To test this hypothesis a cell line in which CD95 signaling was inhibited by overexpression of Bcl-xL was used. In these MCF7-Fas-bcl-xL cells Bcl-xL had no effect on the recruitment of caspase-8 to the DISC. It did not affect the activity of the DISC nor the generation of the caspase-8 active subunits p18 and p10. In contrast, cleavage of a typical substrate for caspase-3-like proteases, poly(ADP-ribose) polymerase, was inhibited in comparison with the control-transfected CD95-sensitive MCF7-Fas cells. To test whether Bcl-xL would inhibit active caspase-8 subunits in the cytoplasm, a number of immunoprecipitation experiments were performed. Using monoclonal antibodies directed against different domains of caspase-8, anti-Bcl-xL antibodies, or fusion proteins of glutathione S-transferase with different domains of caspase-8, no evidence for a direct or indirect physical interaction between caspase-8 and Bcl-xL was found. Moreover, overexpression of Bcl-xL did not inhibit the activity of the caspase-8 active subunits p18/p10. Therefore, in this cell line that has become resistant to CD95-induced apoptosis due to overexpression of Bcl-xL, Bcl-xL acts independently and downstream of caspase-8.
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PMID:Bcl-xL acts downstream of caspase-8 activation by the CD95 death-inducing signaling complex. 945 59

Dietary and synthetic isothiocyanates have cancer chemopreventive activity. Dietary isothiocyanates are formed from glucosinolate precursors of ingested green vegetables. Isothiocyanates are absorbed across intestinal cell membranes by passive diffusion and bind reversibly to plasma protein thiols by thiocarbamoylation. Free isothiocyanate enters cells and is converted to the glutathione conjugate by glutathione S-transferases (GSTs). The glutathione conjugate is exported from cells by multidrug resistance proteins (MRPs), and metabolized in the mercapturic acid pathway to the corresponding mercapturic acid. The isothiocyanate is reformed by fragmentation of mercapturic acid pathway metabolites; it is inactivated by slow hydrolysis to the corresponding amine that is inactive in chemoprevention. Depletion of cellular glutathione and protein thiocarbamoylation activates signal transduction for cancer chemoprevention. Isothiocyanates inhibited and inactivated cytochrome P450 isoforms. They induced increased expression of GST, NADPH: quinone oxidoreductase, aldo-keto reductase and gamma-glutamylcysteine synthetase. These responses were coordinated at the transcription level by nuclear factor-erythroid 2 p45-related factor-2 acting through the antioxidant/electrophile enhancer response element and stimulated by the mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase-1 and c-Jun N-terminal kinase-1 (JNK1) pathway. Isothiocyanates also induced apoptosis of pre-cancerous cells and tumor cells activated by caspase-8 and potentiated by JNK1. The chemopreventive activity of isothiocyanates is influenced by the isothiocyanate bioavailability-as is toxicity, GST polymorphism, protein thiocarbamoylation and probably also by MRP expression. These features of isothiocyanate metabolism and chemoprevention deserve further investigation.
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PMID:Isothiocyanates: mechanism of cancer chemopreventive action. 1198 78

We generated and characterized novel antibodies specific for a cleavage site of human caspase-8/FLICE and its substrate, FLICE-like inhibitory protein (FLIP). The synthetic peptides used as immunogens were CQGDNYQKGIPVETD (#791) and VSEGQLEDSSLLEVD (#1342), which corresponded to cleaved regions of N-terminal fragments of caspase-8 and FLIP generated by active caspase-8, respectively. Each antibody purified from rabbit antiserum reacted specifically with the immunogen but not with the peptide corresponding to the unproteolyzed form, as assessed by ELISA. In vitro cleavage of GST-FLIP by active caspase-8 generated an N-terminal fragment (GST-p43) and a C-terminal one (p12). Consistent with other in vivo data, the FLIP cleavage site follows the Asp residue, LEVD(376)GPAMKNVEF, identified on N-terminal sequencing of the p12 fragment. #1342-antibody (#1342-Ab) recognized the GST-p43 fragment but not the uncleaved protein, thus confirming its specificity. When the antibodies were used for immunoblotting, flow cytometry, and confocal laser microscopy, the proteolysis of caspase-8 and FLIP, and the subcellular localization of their digests could be monitored in apoptotic U937 cells. Interestingly, a significant increase in the percentage of cells exhibiting caspase-8 and FLIP cleavage was observed upon Fas stimulation in interferon-gamma-treated U937 cells, in which the susceptibility to Fas is extremely enhanced. In contrast, U937 cells treated with vitamin D(3) or all-trans retinoic acid showed Fas-resistance, and caspase-8 processing and FLIP cleavage were strongly inhibited. In conclusion, we established a system based on the cleavage site-directed antibodies to monitor the dynamics of caspase-8 processing and activation during apoptosis. Using this system, we found that Fas-susceptibility changes during U937 differentiation occur upstream of caspase-8 processing/activation.
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PMID:Monitoring of caspase-8/FLICE processing and activation upon Fas stimulation with novel antibodies directed against a cleavage site for caspase-8 and its substrate, FLICE-like inhibitory protein (FLIP). 1209 60

There is growing evidence that heavy metals, in general, and mercurial compounds, in particular, are toxic to the human immune system. We have previously shown that methyl mercuric chloride (MeHgCl) is a potent human T-cell apoptogen; moreover, mitochondria appear to be a target organelle for the induction of cell death. The objective of this study was to determine the impact of MeHgCl on mitochondrial function in lymphocytes in terms of modulating reactive oxygen species (ROS) generation, thiol status, and caspase activation. Using the fluorescent probe, 3,3'-dihexyloxacarbocyanine, we demonstrated that exposure to MeHgCl for 1 h resulted in a profound decrease in the mitochondrial transmembrane potential. We next observed the release of cytochrome c from mitochondria into the cytosol; significant translocation was noted between 4 and 8 h following treatment with mercury. ROS generation was monitored by following the conversion of dihydroethidium to the fluorescent product, ethidium. Kinetic analysis indicated that ROS generation was maximal after 16 h of exposure to MeHgCl. The toxicant also depleted the thiol reserves of the cell; glutathione levels were depleted in a dose-dependent fashion reaching minimal levels at 16 h. Real-time RT-PCR analysis demonstrated a significant reduction in both glutathione S-transferase and glutathione peroxidase gene expression in mercury-treated cells. Finally, after 16 h of treatment with MeHgCl, we observed activation of caspase-8, -9, and -3 along with increased expression of caspase-8 and -9. We propose that the target organelle for MeHgCl is the mitochondrion and that induction of oxidative stress is critical to activation of death-signaling pathways. Additonally, mercury acts as a genotoxin significantly altering the expression of genes that affect cell survival and apoptosis.
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PMID:Mercury-induced apoptosis in human lymphocytes: caspase activation is linked to redox status. 1221 6

Hepatoprotection mediated by free radical scavenging molecules such as dimethyl sulfoxide (Me(2)SO) arose the question as to whether this effect involved one or several anti-apoptotic signals. Here, using primary cultures of rat hepatocytes and in vivo thioacetamide-induced liver failure, we showed that Me(2)SO failed to prevent any cleavage of initiator caspase-8 and -9 but constantly inhibited procaspase-3 maturation and apoptosis execution, pointing to an efficient inhibition of cleaved initiator caspase activities. Evidence was recently provided that apoptosis might require both caspase and ASK1/JNK-p38 activities. We demonstrated that this kinase pathway was strongly inhibited in the presence of Me(2)SO whereas overexpression of ASK1 was able to restore caspase-3 activity and apoptosis. Interestingly, we also found that GST M1/2 and GST Alpha1/2 dropped under apoptotic conditions; furthermore transfection of GST M1, A1, or P1 to cells overexpressing ASK1, abolished caspase-3 activity and restored viability. This role of GSTs was further assessed by showing that their high expression level was tightly associated with inhibition of ASK1 activity in Me(2)SO-protected hepatocytes. Together, these results demonstrate that Me(2)SO-mediated hepatoprotection involves a dual inhibition of cleaved initiator caspase and ASK1/JNK-p38 activities. Furthermore, in highlighting the control of apoptosis by GSTs, these data provide new insights for analyzing the complex mechanisms of hepatoprotection.
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PMID:Liver protection from apoptosis requires both blockage of initiator caspase activities and inhibition of ASK1/JNK pathway via glutathione S-transferase regulation. 1237 Jan 86

NF-kappaB regulates the expression of various genes involved in cell growth and differentiation, immune response and inhibition of apoptosis. Recently, some death effector domain (DED)-containing proteins, such as FADD and c-FLIP were reported to activate NF-kappaB. We previously reported that the prodomain-only isoforms of caspase-8 and -10 (PDCasp8/10), containing two DED motifs, could inhibit Fas-mediated apoptosis. Here, we demonstrate that these isoforms also activate NF-kappaB, implying this to be one of the mechanisms by which these polypeptides inhibit apoptosis. The GST pull-down assay revealed that, among upstream kinases that activate NF-kappaB, only NIK and RIP, but not RICK or IKKalpha/beta, could directly bind to PDCasp8/10. In addition, both modules ofDED in PDCasp8/10 were required for these interactions as well as NF-kappaB activation. Experiments using a kinase-dead mutant of IKKalpha and an RIP mutant lacking a kinase domain, both of which function as dominant-negative mutants for their wild-type counterparts, blocked PDCasp8/10-mediated NF-kappaB activation. Using small interfering RNA technology, we further demonstrate that the down-regulation of IKKalpha but not IKKbeta significantly inhibits PDCasp8-mediated NF-kappaB activation. Taken together, these results suggest that caspase-8 and -10 have roles in a non- or anti-apoptotic signaling pathway leading to NF-kappaB activation through RIP, NIK and IKKalpha.
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PMID:Caspase-8 and caspase-10 activate NF-kappaB through RIP, NIK and IKKalpha kinases. 1288 66

Mouse TOSO, the homologue of human TOSO gene, was cloned and characterized in the present study. Using immunofluorescence confocal microscopy we localized TOSO to the cytoplasmic membrane of expressing cells. Using stably transfected mouse TOSO (mTOSO)-expressing Jurkat cells, we show that TOSO protects cells from Fas/Fas ligand- and tumor necrosis factor-induced apoptosis but not from TNF-related apoptosis-inducing ligand-induced apoptosis. The Fas-induced activation of caspase-8 was significantly inhibited by the expression of mTOSO. Using deletion mutants and glutathione S-transferase pull-down approaches, we have shown that mTOSO regulates apoptosis by directly binding to Fas-associated death domain through its C-terminal domain, suggesting the disruption of death-inducing signaling complex formation as mechanism of action. Furthermore, we have expressed mTOSO in transgenic mice and show that mTOSO overexpressing primary T lymphocytes are resistant to Fas/Fas ligand-induced apoptosis.
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PMID:The mouse cell surface protein TOSO regulates Fas/Fas ligand-induced apoptosis through its binding to Fas-associated death domain. 1563 12

The mechanisms involved in regulating mammary cell turnover during the pregnancy-lactation cycle in dairy cows are unclear. The objective of present experiment was to describe expression of genes encoding proteins known to be involved in pathways regulating mammary cell proliferation, apoptosis, differentiation, cell survival, and tissue remodeling. Mammary gland biopsies were taken 7 times during the pregnancy-lactation cycle of 10 dairy cows, and samples were analyzed by immunohistochemistry and real-time PCR. Cell proliferation was greatest during the dry period and apoptosis was high in early dry period and early lactation. Based on Fas (tumor necrosis factor receptor superfamily member 6), Fas ligand, and caspase-3, caspase-8, and caspase-9 gene expression, no indication was found of a stage-dependent shift between the extrinsic and intrinsic pathways leading to apoptosis. Gene expression of microsomal glutathione S-transferase (mGST) did not vary significantly, whereas B-cell leukemia/lymphoma 2 (Bcl-2) and BCL2-associated X protein (Bax) gene expression was greatest during the dry period and early lactation and coincided with high cell turnover. Gene expression of early response genes c-Fos, c-Jun, and c-Myc correlated to neither rate of cell proliferation nor plasma concentration of insulin-like growth factor (IGF)-I and insulin. Gene expression of nuclear factor of kappa light chain gene enhancer in B-cells (NFkappaB) and NFkappaB inhibitor alpha was greatest in the periparturient period, and NFkappaB gene expression coincided with an anticipated need for cell survival factors. Expression of transforming growth factor beta (TGF-beta) receptor 1 and 2 mRNA was greatest in early lactation, whereas TGF-beta1 did not vary significant during the pregnancy-lactation cycle. Even though our results on the TGF-beta system did not comply with other studies, the gene expression pattern of the TGF-beta receptors indicates a role in regulating apoptosis in early lactation. Signal transducer and activator of transcription 5 (STAT5) gene expression was high in the periparturient period, which suggests a role for STAT5 in regulation of mammary cell proliferation and differentiation in dairy cows. Expression of tissue-plasminogen activator, plasminogen activator inhibitor-1, and IGF binding protein 5 genes was greatest in early lactation, suggesting a role for IGF binding protein 5 in coordinating regulation of apoptosis and tissue remodeling.
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PMID:Cellular mechanisms in regulating mammary cell turnover during lactation and dry period in dairy cows. 1848 54

Vitamin A is normally stored in the mammalian liver and is physiologically released depending on the need of the organism for the vitamin. However, there is a compelling evidence showing that even the liver is affected by conditions of high vitamin A intake. Based on these previously reported findings showing negative effects of vitamin A on mammalian tissues, we have investigated the effects of a supplementation with vitamin A at clinical doses (1000-9000 IU/kg day(-1)) on some rat liver parameters. We have analyzed hepatic redox environment, as well as the activity of the mitochondrial electron transfer chain in vitamin A-treated rats. Additionally, activity of the detoxifying enzyme glutathione S-transferase was checked. Also, caspase-3 and caspase-8 and tumor necrosis factor-alpha levels were quantified to assess either cell death or inflammation effects of vitamin A on rat liver. We found increased free radical production and, consequently, increased oxidative damage in biomolecules in the liver of vitamin A-treated rats. Interestingly, we found increased mitochondrial electron transfer chain activity, as well as glutathione-S-transferase enzyme activity. Neither caspases activity, nor tumor necrosis factor-alpha levels change in this experimental model. Our results suggest a pro-oxidant, but not pro-inflammatory effect of vitamin A on rat liver.
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PMID:Evaluation of redox and bioenergetics states in the liver of vitamin A-treated rats. 1932 36

Melanoma differentiation associated gene-7/interleukin-24 (mda-7/IL-24) is a novel cytokine displaying selective apoptosis-inducing activity in transformed cells without harming normal cells. The present studies focused on clarifying the mechanism(s) by which glutathione S-transferase (GST)-MDA-7 altered cell survival of human renal carcinoma cells in vitro. GST-MDA-7 caused plasma membrane clustering of CD95 and the association of CD95 with procaspase-8. GST-MDA-7 lethality was suppressed by inhibition of caspase-8 or by overexpression of short-form cellular FLICE inhibitory protein, but only weakly by inhibition of cathepsin proteases. GST-MDA-7-induced CD95 clustering (and apoptosis) was blocked by knockdown of acidic sphingomyelinase or, to a greater extent, ceramide synthase-6 expression. GST-MDA-7 killing was, in parallel, dependent on inactivation of extracellular signal-regulated kinase 1/2 and on CD95-induced p38 mitogen-activated protein kinase and c-jun NH(2)-terminal kinase-1/2 signaling. Knockdown of CD95 expression abolished GST-MDA-7-induced phosphorylation of protein kinase R-like endoplasmic reticulum kinase. GST-MDA-7 lethality was suppressed by knockout or expression of a dominant negative protein kinase R-like endoplasmic reticulum kinase that correlated with reduced c-jun NH(2)-terminal kinase-1/2 and p38 mitogen-activated protein kinase signaling and maintained extracellular signal-regulated kinase-1/2 phosphorylation. GST-MDA-7 caused vacuolization of LC3 through a mechanism that was largely CD95 dependent and whose formation was suppressed by knockdown of ATG5 expression. Knockdown of ATG5 suppressed GST-MDA-7 toxicity. Our data show that in kidney cancer cells GST-MDA-7 induces ceramide-dependent activation of CD95, which is causal in promoting an endoplasmic reticulum stress response that activates multiple proapoptotic pathways to decrease survival.
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PMID:MDA-7/IL-24-induced cell killing in malignant renal carcinoma cells occurs by a ceramide/CD95/PERK-dependent mechanism. 1941 61


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