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)

Fetal rat dorsal root ganglion neurons (7-8 days in culture) were labeled with [3H]arachidonic acid for 24 h. Stimulation with 10 microM bradykinin (BK) for 30 s resulted in nearly 2-fold increases in levels of radioactive diglyceride and arachidonic acid. A similar result was obtained in the absence of receptor stimulation using the Ca2+ channel agonist BAY K 8644 (10 microM, in the presence of 100 mM potassium chloride) or the Ca2+ ionophore, ionomycin (2.5 microM). If Ca2+ influx was inhibited by adding 3 mM Co2+, a blocker of voltage-sensitive calcium channels, or 2.5 mM EDTA, then BK-stimulated accumulation of both arachidonate and diglyceride was inhibited. These data suggest Ca2+ influx is required for ligand-stimulated accumulation of both arachidonate (a product of diglyceride-lipase or phospholipase A2) and diglyceride (a product of phospholipase C). Two distinct populations of channels may be involved in these reactions since pretreatment with 10 microM nifedipine or 50 microM verapamil (agents which block a subset of voltage-sensitive Ca2+ channels) inhibited BK-stimulated accumulation of arachidonic acid, but did not inhibit diglyceride accumulation. Such functional discrimination appears to have physiological importance; the inhibitory effect of nifedipine and verapamil on BK-stimulated arachidonate release was mimicked by pretreatment with peptides which decrease Ca2+ channel conductance in dorsal root ganglion neurons. The three peptides used were 1 microM neuropeptide Y, 10 microM somatostatin, and 10 microM [N-MePhe3,D-Pro4]-morphiceptin. The effect of neuropeptide Y was blocked by pretreatment with pertussis toxin.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Modulation by neuropeptides of bradykinin-stimulated second messenger release in dorsal root ganglion neurons. 197 11

The phospholipase C (PLC)-mediated hydrolysis of membrane phosphoinositides is an important signal transduction pathway coupled to the cell-surface receptors for several hormones and growth factors. In addition, PLC activity can be modulated by changes in intracellular calcium and activation of GTP binding proteins. In this report, differential activation of PLC in the human keratinocyte cell line SCC-12F was studied as judged by specific patterns of inositol phosphate formation. Several hormones and growth factors previously shown to stimulate PLC in a variety of cell types were screened for activity in SCC-12F cells. Only bradykinin was active, stimulating the PLC-dependent generation of inositol (1,4,5) triphosphate (Ins(1,4,5)P3). Ins(1,4,5)P3 was rapidly metabolized to inositol(1,4)biphosphate (Ins(1,4)P2) and inositol(1,3,4,5)tetrakisphosphate (Ins(1,3,4,5)P4), and subsequently degraded to inositol monophosphates. The response elicited by bradykinin was concentration dependent (EC50 value of 50 nM), suggesting involvement of a specific bradykinin receptor. Treatment of these cells with the calcium ionophore A23187 appeared to result in the direct formation of Ins(1,4)P2 without Ins(1,4,5)P3 as precursor. Treatment of the cells with AIF4-, a putative activator of GTP binding proteins, resulted in the generation of inositol monophosphates as the major metabolites in the absence of detectable Ins(1,4,5)P3 formation. Taken together, these observations suggest that the PLC complex present in SCC-12F cells can be differentially activated to yield either Ins(1,4,5)P3, Ins(1,4)P2, or InsP. The observed effects may be due to a direct PLC-dependent hydrolysis of the appropriate membrane phosphoinositide.
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PMID:Inositol phosphate formation in the human squamous cell carcinoma line SCC-12 F: studies with bradykinin, the calcium ionophore A23187, and sodium fluoride. 198 86

Bradykinin (BK) and phorbol 12-myristate 13-acetate (PMA) both stimulate the hydrolysis of phosphatidylcholine (PC) in human fibroblasts, resulting in the formation of phosphatidic acid (PA) and diacylglycerol (DG) (Van Blitterswijk, W.J., Hilkmann, H., de Widt, J., and Van der Bend, R.L. (1990) J. Biol. Chem. 266, 10337-10343). Stimulation with BK resulted in the rapid and synchronous formation of [3H]choline and [3H]myristoyl-PA from the correspondingly prelabeled PC, indicative of phospholipase D (PLD) activity. In the presence of ethanol or n-butanol, transphosphatidylation by PLD resulted in the formation of [3H]phosphatidylethanol or - butanol, respectively, at the cost of PA and DG formation. This suggests that PC-derived DG is generated via a PLD/PA phosphohydrolase pathway. A more pronounced but delayed formation of these products was observed by PMA stimulation. The Ca2+ ionophore ionomycin also activated PLD and accelerated (synergized) the response to PMA. Both [3H] choline and [3H]phosphocholine were released into the extracellular medium in a time- and stimulus-dependent fashion, without apparent changes in the high intracellular levels of [3H]phosphocholine. The protein kinase C (PKC) inhibitors staurosporin and 1-O-hexadecyl-2-O-methylglycerol inhibited BK- and PMA-induced activation of PLD. Down-regulation of PKC by long-term pretreatment of cells with phorbol ester caused a dramatic drop in background [3H]choline levels, while subsequent stimulation with BK, ionomycin, or PMA failed to increase these levels and failed to induce transphosphatidylation. From these results we conclude that PLD activation is entirely mediated by (downstream of) PKC. Unexpectedly, however, BK stimulation of these PKC-depleted cells caused a marked generation of DG from PC within 15 s, which was not seen in BK-stimulated control cells, suggesting PC breakdown by a phospholipase C (PLCc). We conclude that cells stimulated with BK generate DG via both the PLCc and the PLD/PA hydrolase pathway, whereas PMA stimulates mainly the latter pathway. BK stimulation of normal cells leads to activation of PKC and, by consequence, to attenuation of the level of PLCc-generated DG and to stimulation of the PLD pathway, whereas the reverse occurs in PKC-down-regulated cells.
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PMID:Phospholipid metabolism in bradykinin-stimulated human fibroblasts. II. Phosphatidylcholine breakdown by phospholipases C and D; involvement of protein kinase C. 203 86

Human umbilical vein endothelial cells (HUVECS) were challenged with thrombin in the presence of [3H]acetate to stimulate the production of radiolabeled platelet activating factor (PAF, 1-O-alkyl-2-[3H]acetyl-sn-glycero-3-phosphocholine, 1-O-alkyl-2-[3H]acetyl-GPC). The 3H-product was isolated by thin-layer chromatography, and 1-radyl-2[3H],3- diacetylglycerols were prepared by phospholipase C digestion and subsequent acetylation at the sn-3 position. When the 1-radyl-2[3H],3-diacetylglycerols were analyzed by zonal thin-layer chromatography, 96-97% of the radiolabeled derivative migrated with 1-acyl-2,3-diacetylglycerol standard. Only minor amounts (3-4%) of 1-alkyl-2[3H],3-diacetylglycerol were observed, demonstrating that the predominant acetylated product synthesized by thrombin-stimulated HUVECS was 1-acyl-2-[3H]acetyl-GPC. This relative abundance of 1-acyl-2-[3H]-acetyl-GPC was not significantly affected by thrombin dose, incubation time, or cell passage, and was also observed in HUVECS challenged with ionophore A23187. In addition, the acetylated product from ionophore A23187- or bradykinin-stimulated bovine aortic endothelial cells contained 90% 1-acyl-2-[3H]acetyl-GPC, suggesting that the synthesis of the 1-acyl PAF analog is not unique to HUVECS. These findings demonstrate that PAF is a minor synthetic component of HUVECS and bovine aortic endothelial cells. In light of the integral role which the vascular endothelial cell plays in the regulation of thrombosis, these findings also suggest that the production of 1-acyl-2-acetyl-GPC may be biologically important.
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PMID:Synthesis of 1-acyl-2-[3H]acetyl-SN-glycero-3-phosphocholine, a structural analog of platelet activating factor, by vascular endothelial cells. 203 29

ATP and ADP, in concentrations ranging from 1-100 microM, increased the release of [3H]choline and [3H]phosphorylcholine (P-choline) from bovine aortic endothelial cells (BAEC) prelabelled with [3H]choline. This action was detectable within 5 minutes and was maintained for at least 40 minutes. ATP and ADP were equiactive, and their action was mimicked by their phosphorothioate analogs (ATP gamma S and ADP beta S) and adenosine 5'-(beta, gamma imido) triphosphate (APPNP), but not by AMP, adenosine, and adenosine 5'-(alpha, beta methylene)triphosphate (APCPP): these results are consistent with the involvement of P2Y receptors. ATP also induced an intracellular accumulation of [3H]choline: the intracellular level of [3H]choline was increased 30 seconds after ATP addition and remained elevated for a least 20 minutes. The action of ATP on the release of choline metabolites was reproduced by bradykinin (1 microM), the tumor promoter phorbol 12-myristate 13-acetate (PMA, 50 nM), and the calcium ionophore A23187 (0.5 microM). Down-regulation of protein kinase C, following a 24-hour exposure of endothelial cells to PMA, abolished the effects of PMA and ATP on the release of choline and P-choline, whereas the response to A23187 was maintained. These results suggest that in aortic endothelial cells, ATP produces a sustained activation of a phospholipase D hydrolyzing phosphatidylcholine. The resulting accumulation of phosphatidic acid might have an important role in the modulation of endothelial cell function by adenine nucleotides. Stimulation of phospholipase D appears to involve protein kinase C, activated following the release of diacylglycerol from phosphatidylinositol bisphosphate by a phospholipase C coupled to the P2Y receptors (Pirotton et al., 1987a).
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PMID:Adenine nucleotides modulate phosphatidylcholine metabolism in aortic endothelial cells. 210 83

Several classes of growth factors can be distinguished that act through different signal transduction pathways. One class is constituted by the peptide growth factors that bind to receptors with ligand-dependent protein tyrosine kinase activity. Another class of mitogens activates a phosphoinositide-specific phospholipase C via a receptor-linked G protein. An intriguing member of this class is lysophosphatidic acid (LPA). LPA mitogenicity is not dependent on other mitogens and is blocked by pertussis toxin. LPA evokes at least three separate signalling cascades: (i) activation of a pertussis toxin-insensitive G protein mediating phosphoinositide hydrolysis; (ii) release of arachidonic acid in a GTP-dependent manner, but independent of prior phosphoinositide hydrolysis; and (iii) activation of a pertussis toxin-sensitive Gi protein mediating inhibition of adenylate cyclase. The peptide bradykinin mimics LPA in inducing responses (i) and (ii), but fails to activate Gi and to stimulate DNA synthesis. Our results suggest that the mitogenic action of LPA occurs through Gi or a related pertussis toxin substrate and that, unexpectedly, the phosphoinositide hydrolysis pathway is neither required nor sufficient, by itself, for mitogenesis.
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PMID:Growth factor-like action of lysophosphatidic acid: mitogenic signalling mediated by G proteins. 211 27

Specific prostaglandins have been identified that mediate the sympathetic postganglionic neuron-terminal dependent hyperalgesia induced by bradykinin and norepinephrine, prostaglandin E2 and prostacyclin, respectively. In this study we evaluated the hypothesis that bradykinin and norepinephrine stimulate prostaglandin production in the rat, via distinct phospholipases. We found that, in normal skin, bradykinin hyperalgesia is inhibited by the phospholipase A2 inhibitor, mepacrine, but not by the phospholipase C inhibitor, neomycin and is mimicked by phospholipase A2. In chloroform-treated skin or when co-injected with A23187, bradykinin-induced hyperalgesia was found to consist of two components, one resulting from prostaglandin E2 synthesis (phospholipase A2-dependent) and one resulting from prostacyclin synthesis (phospholipase C-dependent). This latter component is blocked by Quin 2 and verapamil and also inhibited by yohimbine, an alpha 2 receptor antagonist. Arachidonic acid induces a dose-dependent hyperalgesia that was found to be like bradykinin-hyperalgesia in untreated skin (prostaglandin E2-mediated and phospholipase A2-dependent). In chloroform-treated skin or in the presence of A23187, arachidonic acid like bradykinin led to the production of prostacyclin as well as prostaglandin E2. Norepinephrine does not produce hyperalgesia in untreated skin, but in chloroform pretreated skin or in the presence of the calcium ionophore A23187, norepinephrine produces a potent dose-dependent hyperalgesia. This hyperalgesia is prevented by sympathectomy and suppressed by the calcium antagonists Quin 2 and verapamil. It is also suppressed by indomethacin and neomycin but not by SC19220 and mepacrine and is mimicked by phospholipase C.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of distinct phospholipases mediating bradykinin and noradrenaline hyperalgesia. 212 75

The modulatory role of endogenous cellular glycosphingolipids in bradykinin-stimulated myo-inositol 1,4,5-trisphosphate (InsP3) formation by MDCK cells was evaluated utilizing the glucosylceramide synthase inhibitor, threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP). Bradykinin-stimulated InsP3 formation in intact cells and in isolated plasma membranes was significantly enhanced when cells were first depleted of their glucosphingolipids. The effect of glucosphingolipid depletion on phospholipase C activity was dependent on the duration of exposure to the inhibitor and the cellular level of glucosylceramide. Inclusion of glucosylceramide in the culture medium prevented the stimulatory effect of PDMP on InsP3 formation. It is concluded that membrane glucosphingolipids may regulate phospholipase C activity.
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PMID:Glucosphingolipid dependence of hormone-stimulated inositol trisphosphate formation. 214 85

We showed previously that transformation by cytoplasmic and membrane-associated oncogenes including ras results in uncoupling between surface stimulation by platelet-derived growth factor, bombesin, and serum and activation of intracellular phospholipase C (PLC); this uncoupling does not involve alterations at the receptor or effector enzyme levels (T. Alonso, R. O. Morgan, J. C. Marvizon, H. Zarbl, and E. Santos, Proc. Natl. Acad. Sci. USA 85:4271-4275, 1988). In this study, we stimulated normal and oncogene-transformed NIH 3T3 cells with fluoroaluminate (AIF4-), thus directly activating PLC-associated G protein(s) and bypassing the receptor step. A1F4(-)-elicited PLC responses were significantly impaired in transformed cells when compared with those in their normal counterparts, suggesting that the uncoupling of PLC is the result, at least in part, of functional impairment at the G-protein level. Transformation by ras oncogenes has also been reported to result in enhanced PLC response to bradykinin resulting from increased receptor numbers (G. Parries, R. Hoebel, and E. Racker, Proc. Natl. Acad. Sci. USA 84:2648-2652, 1987; J. Downward, J. de Gunzburg, R. Riehl, and R. Weinberg, Proc. Natl. Acad. Sci. USA 85:5774-5778, 1988). We demonstrate here that transformation by other membrane-associated and cytoplasmic oncogenes also results in increased responsiveness to bradykinin ("supercoupling") and enhanced receptor numbers. However, there is no direct correlation between the number of receptors and the enhancement in responsiveness, suggesting that other factors besides receptor number are also involved in the enhanced responses. We propose that a common effect of transformation by cytoplasmic and membrane-associated oncogenes is functional alteration of coupling G proteins and that a similar modification of different kinds of G proteins may account for the pleiotropic alterations of signal transduction (uncoupling and supercoupling) observed.
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PMID:Alterations of G-protein coupling function in phosphoinositide signaling pathways of cells transformed by ras and other membrane-associated and cytoplasmic oncogenes. 216 May 94

The murine BALB/c 3T3 fibroblast clone SV-T2 (3T3 cells) expresses receptors for the nonapeptide bradykinin. In these cells, bradykinin stimulates both inositol phosphate (InsP) formation and arachidonic acid release by independently activating phospholipase C and phospholipase A2, respectively. These actions of bradykinin are mediated by a receptor(s) coupled to pertussis toxin-insensitive guanine nucleotide-binding proteins. Bradykinin-stimulated increases in InsP lead to the mobilization of intracellular Ca2+. We examined the expression of 3T3 receptors for bradykinin in oocytes from Xenopus laevis, cells capable of in vitro expression of foreign mRNA for receptors coupled to the mobilization of Ca2+. Poly(A)+ mRNA was prepared from 3T3 cells and expression of receptors for bradykinin was demonstrated by agonist-mediated stimulation of 45Ca2+ efflux from oocytes injected with 50 ng of poly(A)+ RNA. Bradykinin-stimulated efflux of 45Ca2+ was dose dependent (EC50 = 15 nM) and blocked by the specific mixed B1,B2 bradykinin antagonist NPC 567 but not by the B1 antagonist desArg9[Leu8]bradykinin. Size fractionation of 3T3 poly(A)+ RNA on a sucrose gradient demonstrated a single peak of bradykinin-stimulated 45Ca2+ efflux, with an approximate mRNA size of 4.5 kilobases. Bradykinin-stimulated 45Ca2+ efflux in size-fractionated mRNA was clearly separable from response to [Arg]vasopressin at another receptor linked to InsP formation and Ca2+ mobilization in 3T3 cells.
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PMID:Functional expression of B2 bradykinin receptors from Balb/c cell mRNA in Xenopus oocytes. 216 13


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