Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:4.6.1.1 (adenylate cyclase)
 19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cholera diarrhoea is due to the action of a toxin that acts on all animal cells by stimulating the enzyme adenylate cyclase, which catalyses the production oc cyclic AMP from ATP. In intestinal brush border cells raised cyclic AMP levels result in increased secretion of chloride ions, leading to fluid accumulation in the gut. Escherichia coli produces a similar toxin. The receptor for cholera toxin on the cell membrane appears to be a complex containing the ganglioside GGnSLC (or GM1). Cholera toxin is a protein composed of two different kinds of subunits linked non-covalently. Each toxin molecule has one subunit A and four or more subunits B. Subunit B is inactive but binds to the ganglioside GGnSLC on the cell surface. Subunit A does not bind to cell membranes or gangioside and is slightly toxic to intact cells but strongly and instantly active in lysed cells. The binding of whole toxin through the B subunit to the cell is followed by a lag before subunit A penetrates the cell membrane (leaving subunit B on the surface) and stimulates the adenylate cyclase. The stimulation of adenylate cyclase depends on the presence of NAD and other co-factors present in the cell sap.
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PMID:The nature and action of cholera toxin. 79

Cholera toxin, or peptide A1 from the toxin, activates adenylate cyclase solubilized from rat liver with Lubrol PX, provided that cell sap, NAD+, ATP and thiol-group-containing compounds are present. The activation is abolished by antisera to whole toxin, but not to subunit B.
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PMID:Activation by cholera toxin of adenylate cyclase solubilized from rat liver. 98 19

The toxin of Vibrio Cholera causes fluid secretion from the small intestine by stimulation of adenylate cyclase and elevation of intracellular cyclic AMP concentrations. The toxin is a protein composed of subunits responsible for binding to cell membranes and a subunit responsible for the activation of adenylate cyclase. The binding subunits (B) are non-covalently bonded to the active subunit (A). The latter is composed of two polypeptides A1 and A2 linked by a disulphide bridge. Exposure of the intestine to toxin results in rapid binding to the brush border membrane. Thence follows a gradual increase in adenylate cyclase activity, and stimulation of electrolyte and fluid secretion. Enzyme localization studies show that the brush border does not contain adenylate cyclase. Thus the stimulation of adenylate cyclase by toxin which interacts with the brush border must be indirect. From recent studies it seems that an activator of adenylate cyclase can be found in cytosol from toxin-treated cells. Incubation of toxin with cytosol or dithiothreitol results similarly in the formation of an activator. Preincubation of toxin with cytosol results in more rapid activation of adenylate cyclase in liver cell membranes than direct addition of cytosol and toxin. Preincubation of cholera toxin for activation, by cytosol, is presumed to be due to the splitting of the disulphide band between the A1 and A2 components of the active subunit. The stimulatory ability resides in A1 and both A1 and NAD are required for the activation of adenylate cyclase. The toxin-stimulated adenylate cyclase has similar characteristics to the enzyme stimulated by non-hydrolysable analogs of GTP such as guanylylimidodiphosphate (GppNHp). Stimulation by either cholera or GppNHp is irreversible, the responses to catecholamines are enhanced and the enzyme can be solubilized in the activated state.
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PMID:Sequence of events in the activation of adenylate cyclase by cholera toxin. 100 63

Activation of adenylate [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] by cholera toxin (84,000 daltons, 5.5 S) is demonstrated in plasma membrane fragments of mouse ascites cancer cells. The activation of adenylate cyclase is mediated by a macromolecular cyclase activating factor (MCAF), which has a sedimentation constant of 2.7 S and a molecular weight of about 26,000. MCAF is derived from, and may be identical to the "A fragment" of cholera toxin. Generation of MCAF depends on prior interaction of cholera toxin with either dithiothreitol, NADH, NAD, or a low-molecular-weight component (less than 700 daltons) present in cytoplasm. Subsequent exposure of this pretreated cholera toxin to cell membranes from a variety of mouse ascites cancer cells is followed rapidly by the appearance of MCAF, which no longer requires dithiothreitol, NADH, or NAD for the activation of adenylate cyclase. Activation of adenylate cyclase by MCAF in ascites cancer cell membrane fragments is not reversed by repeated washing of these membrane fragments. Adenylate cyclase in normal cell membrane fragments fails to respond either to cholera toxin or MCAF in the presence of dithiothreitol. In striking contrast, the adenylate cyclase in membrane fragments from five ascites cancer cells responds to either MCAF or native cholera toxin preincubated with dithiothreitol, NADH, or NAD.
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PMID:Cholera toxin activation of adenylate cyclase in cancer cell membrane fragments. 105 74

Similarities exist between the properties of adenylate cyclase after stimulation by cholera toxin and after stimulation by guanylylimidodiphosphate (Gpp-(NH)p). Thus a strong stimulation is achieved by both agents, the stimulation is essentially irreversible, the action of certain hormones is enhanced and the enzyme can be solublized with Lubrol PX in the activated state. Because of these similarities the interaction of cholera toxin and Gpp(NH)p on adenylate cyclase was examined. It was found that prior activation of rat liver adenylate cyclase by cholera toxin in vivo, or by cholera toxin and NAD in homogenates, blocked the stimulatory effect of Gpp(NH)p. Furthermore under conditions in which the effect of Gpp(NH)p was less than that of cholera toxin, inhibition of stimulation by cholera toxin was seen. Stimulation of adenylate cyclase by maximal concentrations of Gpp(NH)p, but not by submaximal concentrations, blocked the stimulatory effect of cholera toxin. The mutant interference of the actions of these two agents suggests a common target in the regulatory mechanism of the adenylate cyclase complex.
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PMID:Effects of cholera toxin on adenylate cyclase. Studies with guanylylimidodiphosphate. 118 56

NG108-15 neuroblastoma x glioma somatic hybrid cells were permeabilized in the presence of [32P]NAD+ and then cultured for 18 h. Resolution of the cell proteins on polyacrylamide gels revealed [32P]ADP-ribosylation of five major protein species with molecular mass values of 52 kDa, 44 kDa, 35 kDa, 30 kDa and 25 kDa. A similar pattern of labelling was also seen when NG108-15 cell membranes were incubated with [32P]NAD+ and hydrolysis of the product revealed mono(ADP-ribosyl)ation. Immunoprecipitation of these products with anti-Gs alpha antiserum revealed a single band identical to cholera toxin substrate. Culture of [32P]NAD(+)-loaded cells for 18 h in the presence of 50 mM-nicotinamide inhibited the eukaryotic mono(ADP-ribosyl)transferase activity. Inhibition of the eukaryotic enzyme was also accompanied by an increase in the abundance of Gs alpha, whether measured by Western blotting with anti-Gs alpha antibody (two separate antisera) or by cholera toxin-dependent [32P]ADP-ribosylation. There was no accompanying change in the abundance of G beta. The increase in Gs alpha abundance in nicotinamide-treated NG108-15 cells was accompanied by a 2-fold increase in basal adenylate cyclase activity (measured in the presence of GTP), and by a smaller but significant increase in iloprost-dependent activation of adenylate cyclase. Receptor number or affinity was not affected by nicotinamide, since this treatment did not alter the binding parameters of [3H]iloprost to NG108-15 cell membranes. Short-term exposure of cells to nicotinamide for 1 h revealed no significant difference in either basal or agonist-stimulated adenylate cyclase activity. These results reveal that mono(ADP-ribosyl)ation of Gs alpha by eukaryotic ADP-ribosyltransferase modifies the abundance and activity of Gs alpha in NG108-15 cells, and hence may play a role in the hormonal regulation of cell function.
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PMID:Gs alpha is a substrate for mono(ADP-ribosyl)transferase of NG108-15 cells. ADP-ribosylation regulates Gs alpha activity and abundance. 128 Jan 14

Transforming growth factor-beta (TGF beta) produced by osteoblasts is present in high levels in bone and influences bone formation, replication of bone cells, and expression of osteoblast protein products. Interactions between bone active hormones and locally released and activated TGF beta were studied by examining the influence of TGF beta preincubation on PTH, calcitonin (CT), and vitamin D receptors in an osteoblastic cell line (UMR 106-06). Preincubation of UMR 106-06 cells with 1 ng/ml TGF beta for 3 days increased specific binding of [125I]PTH-related protein (PTHrP)(1-84) to 140% of that in control cells, but [125I]salmon CT binding decreased to 50% of controls. Binding isotherms indicated that the changes in binding were due to altered receptor numbers since affinities for 125I-labeled PTH and CT remained unchanged. The effect on receptor levels was time dependent, requiring 24 h preincubation with TGF beta for measurable changes, and dose dependent, with maximal effects seen with 1 ng/ml TGF beta. Binding of [3H]1,25(OH)2 vitamin D3 was increased to 130% of control in cytosolic extracts of UMR 106-06 cells pretreated for 3 days with 1 ng/ml TGF beta. Scatchard plots suggested an increase in receptor number without change in affinity. The adenylate cyclase response to PTH increased to 150% of control cells after 3 days of treatment with 1 ng/ml TGF beta; however, the adenylate cyclase response to CT was little changed. Forskolin- and cholera toxin-stimulated adenylate cyclase responses were increased by TGF beta treatment to 130-160% of control, indicating an increase in the stimulatory subunit of the G protein. Increased abundance of both Gs and Gi proteins were indicated by increased cholera toxin- or pertussis toxin-dependent [32P] NAD ribosylation of 47-kilodalton (kDa) and 42-kDa or 40-kDa proteins, respectively, in TGF beta-treated cells. Our data support a complex regulatory effect of TGF beta on UMR 106-06 cells with increases in PTH receptors, vitamin D receptors, and G proteins, whereas there is an apparent down-regulation of CT receptors. TGF beta might induce a more differentiated osteoblast phenotype of these cells, which already express differentiated features such as high alkaline phosphatase activity, PTH and vitamin D receptors, and collagenase production. Since low doses of PTH stimulate bone formation in vivo, TGF beta released or activated at sites of new bone formation might locally modulate PTH activity be allowing increased PTH receptor and postreceptor effectiveness.
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PMID:Transforming growth factor-beta modulates receptor binding of calciotropic hormones and G protein-mediated adenylate cyclase responses in osteoblast-like cells. 132 61

Whole-cell patch-clamp recordings were obtained from light-responsive on-bipolar cells in retinal slices of the dogfish. Inclusion of the A-subunit of pertussis toxin in the patch-pipette solution resulted in an increase in inward current and membrane conductance, and a block of light-evoked currents of on-bipolar cells. The opposite effect was obtained with the A-subunit of cholera toxin, which blocked light responses, and induced an outward current and a decrease in membrane conductance. These actions were NAD+ dependent. The results show that the G-protein(s) linking glutamate receptors to a cGMP cascade in on-bipolar cells possess sites which are ADP-ribosylated by pertussis and cholera toxins, with no homology to the adenylate cyclase system but possibly with a homology to transducin. Furthermore, inclusion of H-7, a kinase inhibitor in the patch-pipette solution, or of a non-hydrolysable ATP analogue (AMP-PNP) had no effect on light responses, membrane conductance or dark current of on-bipolar cells, suggesting that the components of this cGMP cascade are unlikely to be regulated by protein kinases.
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PMID:The glutamate-receptor linked cGMP cascade of retinal on-bipolar cells is pertussis and cholera toxin-sensitive. 134 16

In enzymatically dispersed enriched rat parietal cells we studied the effect of pertussis toxin on prostaglandin E2 (PGE2)- or somatostatin-induced inhibition of H(+)-production. Parietal cells were incubated in parallel in the absence (control cells) and presence of pertussis toxin (250 ng/ml; 4 h). [14C]Aminopyrine accumulation by both pertussis toxin-treated and control cells was used as an indirect measure of H(+)-production after stimulation with either histamine, forskolin or dibutyryl adenosine 3',5'-cyclic monophosphate (dbcAMP) alone and in the presence of PGE2 (10(-9)-10(-7) M) or somatostatin (10(-9)-10(-6) M). PGE2 inhibited histamine- and forskolin-stimulated [14C]aminopyrine accumulation but failed to alter the response to dbcAMP. Somatostatin was less effective and less potent than PGE2 in inhibiting stimulation by histamine or forskolin and reduced the response to dbcAMP. Pertussis toxin completely reversed inhibition by both PGE2 and somatostatin on histamine- and forskolin-stimulated H(+)-production but failed to affect inhibition by somatostatin of the response to dbcAMP. After incubation of crude control cell membranes with [32P]NAD+, pertussis toxin catalysed the incorporation of [32P]adenosine diphosphate (ADP)-ribose into a membrane protein of molecular weight of 41,000, the known molecular weight of the inhibitory subunit of adenylate cyclase (Gi alpha). Pertussis toxin treatment of parietal cells prior to the preparation of crude membranes almost completely prevented subsequent pertussis toxin-catalysed [32P]ADP ribosylation of the 41,000 molecular weight protein.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Pertussis toxin reverses prostaglandin E2- and somatostatin-induced inhibition of rat parietal cell H(+)-production. 135 83

Rat islets express a pertussis toxin sensitive G-protein involved in receptor-mediated inhibition of insulin secretion. This has been assumed previously to represent "G(i)" which couples inhibitory receptors to adenylate cyclase. Incubation of islet G-proteins with 32P-NAD and pertussis toxin resulted in the labelling of a band of molecular weight 40,000. This band was very broad and did not allow resolution of individual components. Incubation of the radiolabelled proteins with an anti-G(o) antiserum resulted in specific immunoprecipitation of a 32P-labelled band. These results demonstrate that the complement of pertussis toxin sensitive G-proteins in rat islets includes G(o).
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PMID:Immunoprecipitation of a pertussis toxin substrate of the G(o) family from rat islets of Langerhans. 135 45


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