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:3.1.4.3 (phospholipase C)
18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effects of pertussis toxin and cholera toxin on early events of T lymphocyte activation were examined in the T lymphocyte cell line, Jurkat. Pertussis toxin treatment of these T cells increased inositol phosphates production and led to increases in intracellular free calcium concentration. These effects were produced by the isolated B (binding) subunit of pertussis toxin, alone. Inositol phosphates production resulting from perturbation of the T cell antigen receptor-CD3 complex by MAb was not affected by pertussis toxin treatment but was markedly inhibited by cholera toxin. This effect of cholera toxin paralleled elevations in cAMP content. However, forskolin, in concentrations equipotent for cAMP production, was a weaker inhibitor of inositol phosphates production. Cholera toxin inhibition of inositol phosphates production did not result from inhibition of baseline incorporation of inositol into phosphoinositide substrates of phospholipase C. These studies underline the complexity of toxin effects on cellular systems and suggest that other approaches will be required to implicate guanine nucleotide-binding regulatory proteins in control of the early events of T lymphocyte activation. However, the data presented here provide a molecular basis for the clinical observations of lymphocytosis and the in vitro observations of lymphocyte mitogenesis after pertussis toxin stimulation.
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PMID:Bacterial toxins affect early events of T lymphocyte activation. 253 43

NIH-3T3 cells transformed by the EJ-ras oncogene display reduced platelet-derived growth factor (PDGF)-stimulated phospholipase C activity as measured by inositol 1,4,5-triphosphate (IP3) synthesis and Ca2+ mobilization. The lack of PDGF-stimulated Ca2+ mobilization in EJ-ras transformed cells is not due to a loss of IP3 sensitivity, because microinjected IP3 elevates intracellular Ca2+. Treatment of EJ-ras transformed cells with cholera toxin or 8-bromo-cyclic AMP, but not pertussis toxin or the beta-subunit of cholera toxin, results in a slight recovery of PDGF-stimulated IP3 synthesis, a marked increase in intracellular Ca2+ mobilization, and an almost complete recovery of prostaglandin E2 biosynthesis. These data suggest that EJ p21-mediated inhibition of PDGF-stimulated intracellular events can be partially and transiently reversed by cyclic AMP.
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PMID:Cyclic AMP can partially restore platelet-derived growth factor-stimulated prostaglandin E2 biosynthesis, and calcium mobilization in EJ-ras-transformed NIH-3T3 cells. 254 Nov 40

F9 mouse teratocarcinoma and PyS-2 cells in culture incubated with monovalent cations in buffered sucrose solution (0.25 M) can secrete as much as 40% of their total lysosomal enzymes into the medium within 30 min. Longer incubation does not lead to further loss of enzyme, suggesting that only a certain fraction of lysosomes is capable of discharge. The simultaneous presence of sucrose and cation, each at the respective optimal concentrations of 0.25 and 0.15 M, is required for lysosomal discharge (i.e. twice isoosmolarity). The cells remain fully viable. Sodium ions are more effective than lithium and potassium ions, whereas amines and divalent cations are less effective. Other sugars including glucose can replace sucrose to varying extents. Secretion is accompanied by a rapid short-lived rise in the level of cAMP. Forskolin as well as agents that activate G protein such as cholera toxin, AlF4-, and vanadate ions also increase the rate of secretion. Sucrose-Na+ stimulation takes place independently of changes in influx or efflux of calcium ions or changes in the levels of extracellular or free intracellular calcium ions. Neomycin, an inhibitor of phospholipase C, has little effect on secretion. Our results suggest that the secretion observed is mediated by a cAMP-dependent mechanism involving G proteins. Calcium ions and phospholipase C appear to play little or no part in the activation process.
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PMID:Stimulated secretion of lysosomal enzymes by cells in culture. 254 92

In cultured foreskin fibroblasts, bradykinin stimulates inositol phosphate generation, arachidonic acid release, and Na+/H+ exchange, with doses of 1-3 nM yielding half-maximal stimulation. Binding of 3H-bradykinin to these cells demonstrates a single receptor site with a Kd of 2.0 nM and a Bmax of 91 fmoles/mg protein. Bradykinin analogs of the B2 type inhibit this binding. GTP synergizes with bradykinin to stimulate phosphatidylinositol turnover in permeabilized fibroblasts and GTP-gamma-S decreases the Bmax of bradykinin binding to fibroblast membranes, indicating that a G-protein couples the receptor to phospholipase C. Pretreatment of fibroblasts with either cholera or pertussis toxin enhances bradykinin stimulation of inositol phosphate accumulation.
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PMID:Coupling of bradykinin receptors to phospholipase C in cultured fibroblasts is mediated by a G-protein. 254 33

Vasopressin V1 receptors were solubilized from rat liver plasma membranes with the detergent lysophosphatidylcholine. [[3H]Arginine]vasopressin (AVP) binding to the solubilized preparations was specific and saturable, with a dissociation constant of 0.6 nM. Cross-linking of [125I]vasopressin to the solubilized fraction, studied by SDS/polyacrylamide-gel-electrophoretic analysis, demonstrated the presence of a 65 kDa band which was specifically labelled with [125I]vasopressin. Specific binding of [3H]AVP to these solubilized receptors was decreased by guanine nucleotides, but not by adenosine 5'-[beta gamma-imido]triphosphate. Addition of vasopressin increased specific binding of 35S-labelled guanosine 5'-[gamma-thio]triphosphate (GTP[35S]) to the solubilized fractions, indicating co-solubilization of GTP-binding protein(s) [G-protein(s)] and vasopressin receptors. The solubilized fraction was insensitive to both cholera- and pertussistoxin treatment. Immunoblotting of the solubilized fraction with antibodies specific for a phosphoinositide-specific phospholipase C (PI-PLC I) demonstrated the presence of a 60 kDa protein. Anti-PI-PLC I antiserum immunoprecipitated solubilized vasopressin-binding sites from rat liver (V1), but not solubilized vasopressin-binding sites from hog kidney (V2). Similar results were obtained with an anti-PI-PLC I IgG affinity column. The solubilized (V1) receptors were enriched by ion-exchange and high-performance gel-filtration liquid chromatography. Vasopressin-binding activity was co-eluted with PI-PLC I and GTP[S]-binding activity on a DEAE-Sepharose column. The major vasopressin- and GTP[35S]-binding activities were co-eluted with PI-PLC I activity at approx. 240 kDa suggesting that vasopressin receptors from rat liver membranes can be solubilized as a complex of receptor-coupler-effector by using the detergent lysophosphatidycholine.
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PMID:Solubilization of rat liver vasopressin receptors as a complex with a guanine-nucleotide-binding protein and phosphoinositide-specific phospholipase C. 254 66

In order to analyze the complex activities of histamine H2 receptor activation on neutrophils, human HL-60 promyelocytic leukemia cells were differentiated into neutrophils by incubation with dimethyl sufoxide, loaded with the Ca2+-sensitive indicator dyes, indo-1 or fura-2, and the levels of intracellular Ca2+ ([Ca2+]i) measured in a fluorescent-activated cell sorter and fluorimeter, respectively. Histamine increased [Ca2+]i in a dose-dependent manner with a half-maximal concentration (EC50) of approximately 10(-6) to 10(-5) M, which exhibited H2 receptor specificity. Prostaglandin E2 and isoproterenol also induced [Ca2+]i mobilization in HL-60 cells, whereas the cell permeable form of cAMP and forskolin failed to increase [Ca2+]i. Since H2-receptor mediated [Ca2+]i mobilization was not inhibited by reducing the concentration of extracellular Ca2+ nor by the addition of Ca2+ channel antagonists, LaCl3 and nifedipine, [Ca2+]i mobilization is due to the release of Ca2+ from intracellular stores. Furthermore, both 10(-4) M histamine and 10(-6) M fMet-Leu-Phe increased the levels of 1,4,5-inositol trisphosphate. However, histamine-induced mobilization of [Ca2+]i was inhibited by cholera toxin but not by pertussis toxin, whereas the action of fMet-Leu-Phe was inhibited by pertussis toxin but not by cholera toxin. These data suggest that H2 receptors on HL-60 cells are coupled to two different cholera toxin-sensitive G-proteins and activate adenylate cyclase and phospholipase C simultaneously.
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PMID:Multiple signaling pathways of histamine H2 receptors. Identification of an H2 receptor-dependent Ca2+ mobilization pathway in human HL-60 promyelocytic leukemia cells. 255 5

The antigen receptors on B lymphocytes, membrane forms of immunoglobulins, transduce signals regulating B cell growth and differentiation by activating a phosphoinositide-specific phospholipase C. In this report, we describe our recent work aimed at understanding this process in greater detail. We have shown that a GTP-binding component is a necessary cofactor in the stimulation of phospholipase C by mIgM. This component has a number of properties in common with the G protein family of receptor-effector coupling components seen in the adenylate cyclase and other signaling systems. For example, analogues of GTP that cannot be hydrolyzed stimulated mIgM-triggered phosphoinositide breakdown, and an analogue of GDP that cannot be converted to GTP inhibited the reactions. Furthermore, aluminum fluoride, which activates known G proteins, also stimulates phosphoinositide breakdown. The G protein that appears to link mIgM to phospholipase C is not one of the well characterized G proteins involved in the regulation of adenylate cyclase or cGMP phosphodiesterase (GS, Gi, and transducin), as judged by its insensitivity to two bacterial toxins that modify these G proteins, cholera toxin and pertussis toxin. Interestingly, analysis of pertussis toxin sensitivity indicates that there are at least 2 distinct G proteins that couple receptors to phospholipase C. For example, the G protein required for chemotactic peptide receptor signaling in neutrophils is sensitive to pertussis toxin, in contrast to the phosphoinositide signaling G protein in B cells. We have also begun to explore the mechanisms by which mIgM signal transduction can be modulated. Stimulation of protein kinase C with phorbol esters or synthetic DG was found to inhibit mIgM-triggered phosphoinositide breakdown. This regulation probably represents a feedback inhibition that would occur with DG produced by phosphoinositide breakdown. Alternatively, there appear to be other signaling pathways that generate DG33, and they could possibly inhibit phosphoinositide breakdown via protein kinase C. This could be an important locus of regulation during B cell activation. For example, other signals could increase or decrease the potency of this feedback inhibition, and thereby adjust the sensitivity of the B cell to antigen. Alternatively, other agents could stimulate protein kinase C directly, or could stimulate another protein kinase which can do the same thing in this regard, and thereby make the B cell insensitive to antigen by preventing antigen receptor signaling.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Signal transduction via the B cell antigen receptor: involvement of a G protein and regulation of signaling. 255 95

The m1 muscarinic acetylcholine receptor gene was transfected into and stably expressed in A9 L cells. The muscarinic receptor agonist, carbachol, stimulated inositol phosphate generation, arachidonic acid release, and cAMP accumulation in these cells. Carbachol stimulated arachidonic acid and inositol phosphate release with similar potencies, while cAMP generation required a higher concentration. Studies were performed to determine if the carbachol-stimulated cAMP accumulation was due to direct coupling of the m1 muscarinic receptor to adenylate cyclase via a GTP binding protein or mediated by other second messengers. Carbachol failed to stimulate adenylate cyclase activity in A9 L cell membranes, whereas prostaglandin E2 did, suggesting indirect stimulation. The phorbol ester, phorbol 12-myristate 13-acetate (PMA), stimulated arachidonic acid release yet inhibited cAMP accumulation in response to carbachol. PMA also inhibited inositol phosphate release in response to carbachol, suggesting that activation of phospholipase C might be involved in cAMP accumulation. PMA did not inhibit prostaglandin E2-, cholera toxin-, or forskolin-stimulated cAMP accumulation. The phospholipase A2 inhibitor eicosatetraenoic acid and the cyclooxygenase inhibitors indomethacin and naproxen had no effect on carbachol-stimulated cAMP accumulation. Carbachol-stimulated cAMP accumulation was inhibited with TMB-8, an inhibitor of intracellular calcium release, and W7, a calmodulin antagonist. These observations suggest that carbachol-stimulated cAMP accumulation does not occur through direct m1 muscarinic receptor coupling or through the release of arachidonic acid and its metabolites, but is mediated through the activation of phospholipase C. The generation of cytosolic calcium via inositol 1,4,5-trisphosphate and subsequent activation of calmodulin by m1 muscarinic receptor stimulation of phospholipase C appears to generate the accumulation of cAMP.
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PMID:A transfected m1 muscarinic acetylcholine receptor stimulates adenylate cyclase via phosphatidylinositol hydrolysis. 255 56

The catalytic subunit of cholera toxin (CT) can chemically modify the alpha polypeptides of certain G-binding proteins and thus alter their function. In order to study the involvement of CT-sensitive G proteins in T cell activation, we have utilized CT in an in vitro system in which purified, resting human peripheral T cells are activated by anti-CD3 antibodies and rIL-2. Perturbation of the TCR/CD3 molecular complex by anti-CD3 antibodies causes changes in membrane phospholipids and induces a rise in cytoplasmic Ca2+. These events, however, are insufficient to allow progression into cellular proliferation and addition of IL-2 is required. Under these conditions, treatment of cells with a low concentration of CT (2 ng/ml) causes a significant inhibition of the anti-CD3-induced calcium event as well as the anti-CD3 plus IL-2-stimulated proliferation. Under our experimental conditions, inhibition of both proliferation and intracellular Ca2+ elevation by CT requires the involvement of the TCR/CD3 complex. This is supported by the observation that the toxin does not inhibit either the proliferation triggered by ionomycin and PMA or the Ca2+ influx induced by the ionophore. These data suggest that in TCR/CD3-mediated T cell activation CT acts at a point between TCR/CD3 perturbation and the generation of intracellular Ca2+. In view of the ability of CT to activate the alpha subunit of the G protein that stimulates adenyl cyclase (G alpha s), it is possible that the effect of CT on T cells is secondary to intracellular elevation of cAMP. However, measurement of cAMP levels both early after CT addition and at later time points, when proliferation is maximal, reveals lack of cyclic nucleotide accumulation. The presented data are consistent with the interpretation that the CT-mediated inhibition is caused by the modification of a G-binding protein that is either directly or indirectly associated with triggering of T cells via the TCR/CD3 molecular complex. The data also suggest that this protein is not G alpha s and it probably represents an as yet unidentified moiety or one of the several G proteins that have been recently described as regulators of phospholipase C activation.
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PMID:Cholera toxin inhibits resting human T cell activation via a cAMP-independent pathway. 255 13

Bradykinin inhibits vasopressin-stimulated water transport in cortical collecting tubular cells. The biochemical mechanism of this effect was explored by means of primary cultures of rabbit cortical collecting tubular cells. Bradykinin was found to produce a rapid release of calcium from intracellular stores, an increase in sn-1,2-diacylglycerol levels, and a fivefold increase in membrane-bound protein kinase C activity, consistent with stimulation of phospholipase C and activation of protein kinase C in rabbit cortical collecting tubular cells. In addition, bradykinin produced a dose-dependent 46% inhibition of vasopressin-stimulated adenosine 3',5'-cyclic monophosphate (cAMP) formation. Pretreatment with the protein kinase C inhibitors, H-7 and staurosporine, reversed the bradykinin-mediated inhibition of vasopressin-stimulated cAMP accumulation. In contrast, pretreatment with either the phospholipase A2 inhibitor, mepacrine, or pertussis toxin did not prevent the inhibitory effect of bradykinin on vasopressin-stimulated cAMP production, suggesting that the effects are not mediated by prostaglandin E2 or activation of a pertussis-toxin sensitive guanine nucleotide regulatory protein (e.g., Gi). Because bradykinin also inhibits isoproterenol-stimulated cAMP formation but does not inhibit either basal-, forskolin-, or cholera toxin-stimulated cAMP accumulation, the site of this inhibition appears to involve the hormone receptor or coupling of the receptor to the stimulatory guanine nucleotide regulatory subunit (Gs). The results demonstrate that bradykinin stimulates phospholipase C leading to activation of protein kinase C, which then inhibits vasopressin-stimulated cAMP production at the level of the hormone receptor or coupling of the receptor to Gs in cultured cortical collecting tubular cells.
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PMID:Bradykinin activates protein kinase C in cultured cortical collecting tubular cells. 255 39


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