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)

Bradykinin (BK) induced a biphasic increase in 1,2-diacylglycerol (DAG) in both K-ras-transformed fibroblasts (DT) and the parent NIH-3T3 cells. The first phase was coincident with the increase in Ins(1,4,5)P3 resulting from PtdIns(4,5)P2 hydrolysis, and the second, sustained, phase was derived from phosphatidylcholine (PtdCho) hydrolysis. In NIH-3T3 cells, stimulation by BK induced greater production of choline than phosphocholine in [3H]choline-labelled cells and appreciable phosphatidylethanol (PtdEtOH) formation in [3H]myristic acid-labelled cells, suggesting that PtdCho was hydrolysed mainly by a phospholipase D (PLD) activity. Pretreatment with propranolol, an inhibitor of phosphatidate phosphohydrolase, markedly diminished the second DAG accumulation, supporting the above notion. In DT cells, BK induced predominantly phosphocholine generation and little PtdEtOH formation, indicating that the PtdCho hydrolysis was due to a phospholipase C (PLC) activity. The BK-induced oscillations in intracellular Ca2+ concentration ([Ca2+]i) observed in single DT cells [Fu, Sugimoto, Oki, Murakami, Okano & Nozawa (1991) FEBS Lett. 281, 263-266] were detected as a sustained [Ca2+]i elevation when assayed in a cell suspension. A receptor-operated Ca2+ channel blocker, SK&F 96365, suppressed both the BK-induced phosphocholine generation and the sustained [Ca2+]i elevation in a similar dose-dependent manner. These results thus suggested that oscillations in [Ca2+]i are involved in the activation of PtdCho-specific PLC in DT cells.
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PMID:Differential pathways (phospholipase C and phospholipase D) of bradykinin-induced biphasic 1,2-diacylglycerol formation in non-transformed and K-ras-transformed NIH-3T3 fibroblasts. Involvement of intracellular Ca2+ oscillations in phosphatidylcholine breakdown. 157 79

Human preimplantation embryos secrete platelet-activating factor (PAF), which stimulates prostaglandin E2 synthesis from secretory endometrium. This study investigated the action of PAF on phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2)-specific phospholipase C activity in human endometrium. Slices of normal endometrium were incubated with 5 microCi/ml myo-[2-3H] inositol for 3 h at 37 degrees C in 95% O2 and 5% CO2 to label tissue phosphoinositides. Inositol phosphates were extracted using trichloroacetic acid precipitation and diethylether neutralization and production was measured using Dowex 1-X8 anion-exchange column chromatography. PAF induced rapid and concentration-dependent accumulation of inositol phosphates (IP) from secretory endometrium, but had no effect on endometrium removed in the proliferative phase of the menstrual cycle. The IP3 fraction was significantly elevated from a median value of 14.0 c.p.m. mg-1 dry wt [range: 8-41 c.p.m. mg-1 dry wt] to 28.0 c.p.m. mg-1 dry wt [range: 11-87 c.p.m. mg-1 dry wt, P less than 0.002] following 1 min exposure of secretory endometrium to PAF-acether, in the presence of 10 mM LiCl. PAF-induced hydrolysis of PtdIns(4,5)P2 was inhibited by the specific PAF receptor antagonist WEB 2086, in a dose-dependent manner (P less than 0.02), indicating that in human endometrium PtdIns(4,5)P2 hydrolysis is mediated via a PAF receptor. These results indicate that PAF receptor coupling activates endometrial PtdIns(4,5)P2-specific phospholipase C only in the secretory phase of the menstrual cycle, suggesting that the PAF response may be under ovarian steroid regulation. It is proposed that the ability of the endometrium to respond to PAF appears to be a feature of the preparation of this tissue for implantation and that the second messengers generated may play a role in cellular processes involved in the maternal recognition of very early human pregnancy.
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PMID:Platelet-activating factor stimulates phospholipase C activity in human endometrium. 161 19

The activity of a phospholipase C which hydrolyses exogenous phosphatidylinositol-4,5-bisphosphate [( 3H]PtdIns(4,5)P2) in membranes prepared from frozen postmortem human brain and rat brain was investigated. Enzyme characteristics were essentially similar in membranes prepared from frozen postmortem brain and fresh or frozen rat brain. The [3H]PtdIns(4,5)P2 solubilization and assay procedure employed resulted in an efficient availability of the substrate for the enzyme. The non-hydrolysable guanosine triphosphate analogue guanosine 5'-[beta gamma-imido]diphosphate (Gpp[NH]p) stimulated hydrolysis rapidly with a half maximum activity of approximately 25 microM. This stimulation was not specific for guanine nucleotides as ATP, imidodiphosphate and pyrophosphate also caused enzyme activation. However these activation effects could be distinguished by the polyanion spermine. The non-hydrolysable guanine dinucleotide analogue guanosine 5'-[beta-thio]diphosphate acted as a partial agonist thereby inhibiting the stimulatory effect of Gpp[NH]p. Gpp[NH]p-stimulated enzyme activity showed a maximum response in the presence of 1 mM deoxycholate and displayed a pH optima in the range 7.0-7.5. PtdIns(4,5)P2 hydrolysis was observed in the absence of added calcium, but hydrolytic cleavage was inhibited in the presence of divalent ion chelators. Magnesium inhibited PtdIns(4,5)P2 hydrolysis in a concentration-dependent manner. Elucidation of these aspects of the phosphatidylinositol cycle in normal human postmortem brain will permit comparative studies in CNS disease states.
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PMID:Assay of a phosphatidylinositol bisphosphate phospholipase C activity in postmortem human brain. 164 35

Microsomes were prepared from cultured neonatal rat cardiomyocytes. Incubation of microsomes in buffer containing 5 microM CaCl2, 5 mM cholate and 100 nM [3H-]Phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5) P2) resulted in the formation of [3H-]InsP3. GTP-gamma-S (125 microM) stimulated the production of [3H-]InsP3. Microsomes prepared from phorbol ester-treated (100 nM phorbol 12-myristate 13-acetate, PMA) cardiomyocytes showed decreased activities of basal as well as GTP-gamma-S-stimulated [3H-]PtdIns(4,5)P2 hydrolysis. In the microsomes a 15 kD protein was demonstrated to be the major substrate phosphorylated by intrinsic protein kinase C, which was activated by 0.5 mM Ca2+. Addition of phorbol ester (100 nM PMA) enhanced the 32P-incorporation into the 15 kD protein. Protein kinase C, purified from rat brain, in the presence of Ca2+, diglyceride, and phosphatidylserine did not change the phosphorylation pattern any further. In conclusion, it was shown that phorbol ester pretreatment of neonatal rat cardiomyocytes reduces microsomal GTP-gamma-S-stimulated PtdIns(4,5)P2-specific phospholipase C activity, as estimated with exogenous substrate, and that in cardiomyocyte microsomes phorbol ester activates protein kinase C-induced 15 kD protein phosphorylation. The results indicate that phorbol ester may down-regulate alpha 1-adrenoceptor mediated PtdIns(4,5)P2 hydrolysis by activation of protein kinase C-induced 15 kD protein phosphorylation.
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PMID:Phorbol ester and the actions of phosphatidylinositol 4,5-bisphosphate specific phospholipase C and protein kinase C in microsomes prepared from cultured cardiomyocytes. 165 1

Recent studies have shown that the receptor for epidermal growth factor (EGF) can associate with and tyrosine-phosphorylate the gamma-isozyme of phosphoinositide (PtdIns)-specific phospholipase C (PLC gamma), suggesting a possible mechanism for activation of PtdIns hydrolysis by EGF. In the present study, the coupling between PtdIns hydrolysis and PLC gamma tyrosine phosphorylation in WB liver epithelial cells was examined. Peak levels of [P-Tyr]PLC gamma, measured by anti-P-Tyr immunoblotting, occurred at 0.5-2 min of EGF treatment and coincided with the onset of [3H]inositol phosphate production. The termination of PtdIns hydrolysis after EGF stimulation was accompanied by return of [P-Tyr]PLC gamma to near-basal levels. Activation of protein kinase C (PKC) with a phorbol ester inhibited (IC50 = 3-10 nM) both EGF-dependent PtdIns hydrolysis and PLC gamma phosphorylation by more than 90%. Both EGF-stimulated responses were potentiated in cells depleted of PKC by prolonged phorbol ester treatment. At physiological ionic strength, monoclonal antibodies to PLC gamma specifically precipitated (in addition to PLC gamma) the EGF receptor and at least six other [P-Tyr]proteins from extracts of EGF-treated cells. PKC activation had differential effects on the tyrosine phosphorylation of these coprecipitating proteins, i.e. the relative abundance of certain [P-Tyr] proteins decreased, whereas that of another protein increased. In conclusion, EGF-stimulated tyrosine phosphorylation of PLC gamma is broadly correlated with stimulation of PtdIns hydrolysis, consistent with a role for tyrosine phosphorylation in PLC activation. The attendant diacylglycerol release and activation of PKC may terminate PLC gamma activation, in part by inhibiting PLC gamma phosphorylation by the EGF receptor. Our results suggest further that PKC may exert regulatory effects by altering the relationship of PLC gamma to its associated [P-Tyr]proteins.
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PMID:Protein kinase C inhibits epidermal growth factor-dependent tyrosine phosphorylation of phospholipase C gamma and activation of phosphoinositide hydrolysis. 169 45

Endothelin-1 (ET-1)- and lysophosphatidic acid (LPA)-stimulated PtdIns(4,5)P2 hydrolysis has been studied in Rat-1 fibroblasts. Although both agonists caused the dose-dependent accumulation of inositol phosphates, a number of differences were observed. LPA induced a transient increase in Ins(1,4,5)P3 mass which returned to basal levels within 90 s, whereas the response to ET-1 did not desensitize, with levels remaining at 3-4 times basal values for up to 15 min. Stimulated decreases in mass levels of PtdIns(4,5)P2 mirrored Ins(1,4,5)P3 formation for both agonists. Experiments with electropermeabilized cells demonstrated that the effects of both agonists are stimulated by a phospholipase C controlled by a guanine-nucleotide-binding regulatory protein; however, there are differences in the nature of these interactions. The inositol phosphate response to ET-1 is poorly potentiated by guanosine 5'-[gamma-thio]triphosphate (GTP[S]) and markedly inhibited by guanosine 5'-[beta-thio]diphosphate (GDP[S]), whereas that to LPA is potentiated by GTP[S] but is relatively insensitive to GDP[S]. In addition, LPA decreased the lag time for the onset of GTP[S]-stimulated [3H]InsP3 accumulation, whereas ET-1 was without effect. Phorbol 12-myristate 13-acetate treatment of the cells inhibited LPA-stimulated, but not ET-1-stimulated, inositol phosphate formation in both intact and permeabilized cells, suggesting that the site of protein kinase C-mediated phosphorylation may be blocked in ET-1-stimulated Rat-1 cells. The results indicate that the receptor-G-protein-phospholipase C interaction for the two agonists may not conform to the same model.
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PMID:Differences in the regulation of endothelin-1- and lysophosphatidic-acid-stimulated Ins(1,4,5)P3 formation in rat-1 fibroblasts. 176 24

We have previously shown that two ectoenzymes, acetylcholinesterase (AChE) and alkaline phosphatase, are released from the surface and from particulate fractions of the parasite Schistosoma mansoni, by a phosphatidylinositol-specific phospholipase C (PtdIns-PLC) of bacterial origin. Exposure to PtdIns-PLC not only removes large amounts of AChE from the surface of intact, viable Schistosoma in culture, but is accompanied by a concomitant increase in overall levels of AChE in the parasite. The same phenomenon is observed with PtdIns-PLC from two different bacterial sources; Staphylococcus aureus and Bacillus thuringiensis. The increase in AChE levels may be ascribed to de novo synthesis since exposure to PtdIns-PLC, in the presence of the protein-synthesis inhibitor cycloheximide, totally blocked the increase in AChE activity. Furthermore, PtdIns-PLC induced an increased incorporation of [35S]methionine into the AChE immunoprecipitated by a specific anti-AChE serum. This increase is selective for AChE, since total protein synthesis remained almost unchanged after PtdIns-PLC addition, and little or no effect was observed on the enzymatic activity of alkaline phosphatase, which is also glycophosphatidylinositol anchored. Since cleavage of the phosphatidylinositol anchor by PtdIns-PLC should liberate diacylglycerol, which may act as second messenger, we investigated the effect of exogenous diacylglycerols on the synthesis of AChE in S. mansoni. Three different diacylglycerols were tested as possible inducers of AChE activity in the parasite. Both 1-oleoyl-2-acetyl-sn-glycerol and 1,2-dimyristoyl-sn-glycerol were able to increase AChE activity by 35-40% at concentrations of 25 micrograms/ml. A higher concentration of 1,2-dioctanoyl-sn-glycerol (70 micrograms/ml) was needed to produce an equivalent effect. Moreover, addition of phorbol-12-myristate-13-acetate, together with the calcium ionophore A23187, produced a similar increase in AChE activity. Finally, polymixin B, a specific inhibitor of protein kinase C, partially blocked the increase in AChE activity induced by PtdIns-PLC. Our results suggest the involvement of glycophosphatidyl membrane-anchor breakdown products as putative second messengers in the parasite S. mansoni.
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PMID:Phosphatidylinositol-specific phospholipase C induces biosynthesis of acetylcholinesterase via diacylglycerol in Schistosoma mansoni. 184 73

Sequence analysis of a inositol-phospholipid-specific phospholipase C (PtdIns-PLC) purified from bovine brain has led to the isolation of a novel cDNA that encodes this protein. While this cDNA contains two introns, these appear to be removed upon transfection of the cDNA into COS-1 cells. The protein transiently expressed in COS-1 cells shows phosphatidylinositol 4,5-bisphosphate hydrolysing activity which distributes preferentially into the particulate fraction. Comparison of the predicted amino acid sequence of this PtdIns-PLC with other known PtdIns-PLCs reveals a high degree of similarity, throughout all of its sequence, with PtdIns-PLC delta. Thus, we believe that the identification of this cDNA represents evidence for multiple functional-gene products within the delta subclass of PtdIns-PLCs.
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PMID:A second gene product of the inositol-phospholipid-specific phospholipase C delta subclass. 184 83

Using phosphatidylinositol-glycan (PtdIns-glycan) anchored acetylcholinesterase from bovine erythrocytes as substrate, we found PtdIns-glycan-anchor-degrading activity in rat liver and serum [corrected]. The hepatic enzyme was only soluble in detergents, whereas the serum enzyme occurs as soluble, slightly amphiphilic protein. Using 3-trifluoromethyl-3-(m- [125I]iodophenyl)diazirine-labelled acetylcholinesterase as substrate, we showed that the hepatic anchor-degrading enzyme had a cleavage specificity of a phospholipase C, whereas the serum enzyme was a phospholipase D. Both enzyme exhibited maximal activity in slightly acidic conditions and at low ionic strength. They had a high affinity for the PtdIns-glycan anchor of the substrate (Km = 0.1 microM and 0.16 microM, respectively). Both hepatic PtdIns-glycan-specific phospholipase C and serum PtdIns-glycan-specific phospholipase D gave a large increase in activity between 0.1-10 microM Ca2+, indicating that PtdIns-glycan-specific phospholipases are only marginally active at physiological intracellular Ca2+ concentrations. The enzymes were inhibited by heavy metal chelating agents such as 1,10-phenanthroline and 2,2'-bipyridyl but not by the corresponding Fe2+ complexes or non-chelating analogues, indicating that they both require a heavy metal ion for the expression of catalytic activity in addition to Ca2+. Another interesting property of PtdIns-glycan-specific phospholipases is their inactivation by bicarbonate and cyanate. The inactivation was time- and pH-dependent and could be reversed by dialysis. These observations are in agreement with a covalent modification of the enzymes by carbamoylation.
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PMID:Enzymatic properties of phosphatidylinositol-glycan-specific phospholipase C from rat liver and phosphatidylinositol-glycan-specific phospholipase D from rat serum. 184 23

Aluminum (Al) is believed to exert a primary role in the neurotoxicity associated with dialysis encephalopathy and has been suggested to be involved in a number of other neurological disorders, including Alzheimer's disease. Al, complexed with fluoride to form fluoroaluminate (AlF4-), can activate the GTP-binding (G) proteins of the adenylate cyclase and retinal cyclic GMP phosphodiesterase systems. Since an involvement of G-proteins with cerebral phosphoinositide (PtdIns) metabolism has also been suggested, in this study we investigated the interaction of the stable GTP analogue GTP(S), Al salts and NaF with this system. In rat cerebral cortical membranes, GTP(S) dose-dependently stimulated [3H]inositol phosphates ([3H]InsPs) accumulation. This effect was potentiated by carbachol and was partially prevented by the GTP-binding antagonist GDP(S), indicating that CNS muscarinic receptor activation is coupled to PtdIns hydrolysis via putative G-protein(s). GTP(S) stimulation was also inhibited by phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C, which is known to exert a negative feedback control on agonist-stimulated PtdIns metabolism. Both Al salts and NaF mimicked the action of GTP(S) in stimulating PtdIns turnover. Their actions were highly synergistic, suggesting that AlF4- could be the active stimulatory species. However, the stimulatory effects of AlCl3 and/or NaF were not potentiated by carbachol and were not inhibited by GDP(S) and PMA, suggesting that separate sites of action might exist for GTP(S) and AlF4-. In the nervous tissue, activation of PtdIns hydrolysis by Al (probably as AlF4-) may be mediated by activating a regulatory G-protein at a location distinct from the GTP-binding site or by a direct stimulation of phospholipase C.
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PMID:Interaction of aluminum ions with phosphoinositide metabolism in rat cerebral cortical membranes. 194 39


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