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)

We have investigated the stoichiometric relationship of proteins involved in beta-adrenergic-receptor-mediated signal transduction in isolated rat cardiac myocytes. These cells contain about 2.1 x 10(5) beta-adrenergic receptors per cell, as determined by radio-ligand-binding assays. We have assessed the amount of Gs alpha present in myocyte membranes by immunoblotting using a purified glutathione S-transferase-Gs alpha fusion protein as a standard for quantification. By this method, we determined that cardiac myocytes contain about 35 x 10(6) and 12 x 10(6) molecules per cell of the 45 and 52 kDa forms of Gs alpha, respectively. [3H]Forskolin binding assays were used to assess the formation of high-affinity forskolin binding sites representing Gs alpha-adenylate cyclase complexes occurring in response to Gs alpha activation. Quantification of the adenylate cyclase complexes was facilitated by the permeabilization of cells with saponin. The addition of isoprenaline (isoproterenol) and guanosine 5'-[gamma-thio]trisphosphate to saponin-permeabilized myocytes results in the formation of 6 x 10(5) Gs alpha-adenylate cyclase complexes. Taken together, the data presented here demonstrate that, in a physiologically relevant setting, G-protein is present in large stoichiometric excess relative to both receptor and effector. In addition, we show that, overall, only modest signal amplification occurs between receptor and adenylate cyclase. Thus adenylate cyclase (rather than Gs) is the component distal to receptor that limits agonist-mediated increases in cyclic AMP production. Although limited data are as yet available for other G-protein-regulated effectors, we hypothesize that the stoichiometry of signalling components and the extent of signal amplification described for the beta-adrenergic response pathway will be applicable to other G-protein-coupled hormone receptor systems.
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PMID:Quantification of signalling components and amplification in the beta-adrenergic-receptor-adenylate cyclase pathway in isolated adult rat ventricular myocytes. 757 83

Shear stress causes the platelet glycoprotein (Gp) Ib/IX/V to bind to von Willebrand factor, resulting in platelet adhesion. GpIb/IX/V also functions to stimulate transmembranous signaling, leading to platelet activation and the expression of a ligand-receptive GpIIb-IIIa complex. The highly conserved cytoplasmic domain of GpIbalpha binds directly to a dimeric 14-3-3 adapter protein zeta isoform. To explore structural determinants of GpIb/IX/V binding to 14-3-3zeta, the authors examined 14-3-3zeta interactions with GpIbalpha and GpIbbeta in heterologous cells and platelets. Truncations of GpIbalpha at amino acid 542 or 594, or deletions of residues 542 through 590, inhibited binding of 14-3-3zeta. Deletion of GpIbalpha from Trp(570) to Ser(590) eliminated 14-3-3zeta binding, and deletion of the sequence from Arg(542)-Trp(570) enhanced binding of 14-3-3zeta to GpIbalpha. All GpIbalpha mutations that eliminated GpIbalpha binding to the GST-14-3-3zeta fusion protein also eliminated GpIbbeta binding to the fusion protein. Forskolin treatment of Chinese hamster ovary cells expressing wild-type GpIbalpha/beta/IX resulted in the phosphorylation of GpIbbeta associated with enhanced binding of GpIbbeta to GST-14-3-3zeta fusion protein and increased 14-3-3zeta coimmunoprecipitated with GpIbalpha. When intact human platelets aggregated in response to 90 dynes/cm(2) shear stress, 14-3-3zeta disassociated from GpIbalpha. Prostacyclin treatment of platelets inhibited shear stress-induced aggregation and the release of 14-3-3zeta from GpIbalpha. These data demonstrate that amino acid residues in the cytoskeletal interaction domains of GpIbalpha regulate 14-3-3zeta binding to GpIbalpha/beta/IX, and suggest that protein kinase A-dependent phosphorylation of GpIbbeta enhances 14-3-3zeta binding to the GpIb/IX/V complex in human platelets. (Blood. 2000;95:551-557)
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PMID:Cytoplasmic domains of GpIbalpha and GpIbbeta regulate 14-3-3zeta binding to GpIb/IX/V. 1062 61

Despite its key role in potassium homeostasis, transcriptional control of the H(+)-K(+)-ATPase alpha(2)-subunit (HKalpha(2)) gene in the collecting duct remains poorly characterized. cAMP increases H(+)-K(+)-ATPase activity in the collecting duct, but its role in activating HKalpha(2) transcription has not been explored. Previously, we demonstrated that the proximal 177 bp of the HKalpha(2) promoter confers basal collecting duct-selective expression. This region contains several potential cAMP/Ca(2+)-responsive elements (CRE). Accordingly, we examined the participation of CRE-binding protein (CREB) in HKalpha(2) transcriptional control in murine inner medullary collecting duct (mIMCD)-3 cells. Forskolin and vasopressin induced HKalpha(2) mRNA levels, and CREB overexpression stimulated the activity of HKalpha(2) promoter-luciferase constructs. Serial deletion analysis revealed that CREB inducibility was retained in a construct containing the proximal 100 bp of the HKalpha(2) promoter. In contrast, expression of a dominant negative inhibitor (A-CREB) resulted in 60% lower HKalpha(2) promoter-luciferase activity, suggesting that constitutive CREB participates in basal HKalpha(2) transcriptional activity. A constitutively active CREB mutant (CREB-VP16) strongly induced HKalpha(2) promoter-luciferase activity, whereas overexpression of CREBdLZ-VP16, which lacks the CREB DNA-binding domain, abolished this activation. In vitro DNase I footprinting and gel shift/supershift analysis of the proximal promoter with recombinant glutathione S-transferase (GST)-CREB-1 and mIMCD-3 cell nuclear extracts revealed sequence-specific DNA-CREB-1 complexes at -86/-60. Mutation at three CRE-like sequences within this region abolished CREB-1 DNA-binding activity and abrogated CREB-VP16 trans-activation of the HKalpha(2) promoter. In contrast, mutation of the neighboring -104/-94 kappabeta element did not alter CREB-VP16 trans-activation of the HKalpha(2) promoter. Thus CREB-1, binding to one or more CRE-like elements in the -86/-60 region, trans-activates the HKalpha(2) gene and may represent an important link between rapid and delayed effects of cAMP on HKalpha(2) activity.
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PMID:CREB trans-activates the murine H(+)-K(+)-ATPase alpha(2)-subunit gene. 1516 20

The mechanisms by which organotin compounds produce modulations of the endocrine systems and other biological responses are not fully understood. In this study, juvenile salmon were force-fed diet containing TBT (0: solvent control, 0.1, 1 and 10mg/kg fish) for 72 h. Subsequently, fish exposed to solvent control and 10mg TBT were exposed to waterborne concentration (200 microg/l) of the adenylate cyclase (AC) stimulator, forskolin for 2 and 4h. The overall aim of the study was to explore whether TBT endocrine disruptive effects involve second messenger activation. Liver was sampled from individual fish (n=8) at the end of the exposures. The transcription patterns of peroxisome proliferator-activated receptor (PPAR) isotype and acyl-coenzyme A oxidase 1 (ACOX1), aromatase isoform, estrogen receptor-alpha (ER alpha), pregnane X receptor (PXR), CYP3A and glutathione S-transferase (GST) genes were measured by quantitative polymerase chain reaction (qPCR). Our data showed a consistent increase in PPAR alpha, PPAR beta and PPAR gamma mRNA and protein expression after TBT exposure that were inversely correlated with ACOX1 mRNA levels. Forskolin produced PPAR isotype-specific mRNA and protein effects that were modulated by TBT. ACOX1 expression was decreased (at 2h) and increased (at 4h) by forskolin and the presence of TBT potentiated these effects. TBT apparently increased mRNA and protein levels of cyp19a, compared to the solvent control, whereas cyp19b mRNA levels were unaffected by TBT treatment. Combined TBT and forskolin exposure produced respective decrease and increase of mRNA levels of cyp19a and cyp19b, compared with control. TBT decreased ER alpha mRNA at low dose (1mg/kg) and forskolin exposure alone produced a consistent decrease of ER alpha mRNA levels that were not affected by the presence of TBT. Interestingly, PXR and CYP3A mRNA levels were differentially affected, either decreased or increased, after exposure to TBT and forskolin, singly and also in combination. GST mRNA was increased by TBT exposure. Exposure to forskolin alone increased GST expression with time, and combined exposure with TBT potentiated these respective effects. Overall, the present study demonstrates multiple biological effects of TBT given singly or in combination with cAMP activator. There are no studies known to us that have evaluated the endocrine disruptive effects of TBT in the presence of a second messenger activator, and our data suggest that TBT may exert endocrine, biotransformation and lipid peroxidative effects through modulation of cAMP/PKA second messenger signaling with overt physiological consequences.
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PMID:Peroxisome proliferator-activated receptors, estrogenic responses and biotransformation system in the liver of salmon exposed to tributyltin and second messenger activator. 2046 41