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 hydrolysis by phospholipase C from B. cereus of several lecithins of different fatty acyl chain length was examined. The enzyme showed significant activity towards mono-molecularly dispersed short chain lecithins and the reaction obeyed normal Michaelis-Menten kinetics. Rate vs. substrate concentration curves obtained with dihexanoyl-, diheptanoyl- and dioctanoyllecithins showed marked discontinuities in the region of the known critical micelle concentrations for these substrates and distinctly higher rates were obtained just above these levels. Using these three lecithins at levels below their respective critical micelle concentrations, rate increases were noted if the reactions were allowed to proceed to a sufficiently great extent. The presence of deoxycholate in the reaction system had little or no effect on the rate of enzyme-catalysed hydrolysis of lecithins of fatty acyl chain length less than or equal to Cbeta, but for fatty acyl chain lengths greater than C10, significant rate increases occurred. The pH profile for the enzyme activity was also examined.
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PMID:Phospholipase C from Bacillus cereus. Action on some artificial lecithins. 1 20

The alpha 2-C10 adrenergic receptor from human platelets was expressed permanently in Rat-1 fibroblasts. A series of clones that varied in expression of the receptor from 0 to 3.5 pmol/mg of membrane protein were isolated. We have demonstrated recently in cells of one of these clones (1C) that the alpha 2-C10 receptor interacts directly with two distinct pertussis toxin-sensitive G-proteins, Gi2 and Gi3 (Milligan, G., Carr, C., Gould, G. W., Mullaney, I., and Lavan, B.E. (1991) J. Biol. Chem. 266, 6447-6455). High affinity GTPase activity in membranes of cells from the various clones was stimulated by the addition of the alpha 2-adrenergic agonist UK14304, defining that the receptor coupled productively to the G-protein signaling system. Maximal stimulation of high affinity GTPase activity correlated with the levels of receptor expressed. Clones expressing the receptor also demonstrated agonist-mediated inhibition of adenylylcyclase. Futhermore, the alpha 2-C10 receptor in one clone (1C), but not other clones, promoted a marked stimulation in the generation of water-soluble products derived from phosphatidylcholine. The concentration of UK14304 required to produce half-maximal regulation of GTPase activity (20-30 nM), of forskolin-amplified adenylylcyclase activity (30-40 nM), and of choline generation (30-40 nM) were similar. Transphosphatidylation experiments with cells of clone 1C indicated that the receptor-mediated hydrolysis of phosphatidylcholine was via the action of a phospholipase D. All of these effects were attenuated by pretreatment of the cells with pertussis toxin. Dose-effect curves of pertussis toxin-treatment demonstrated similar effective concentrations of the toxin in causing endogenous ADP-ribosylation of both Gi2 and Gi3, inhibition of receptor-stimulated GTPase activity, and phospholipase D activity. Receptor activation of phospholipase D activity was not dependent upon prior phospholipase C-dependent activation of protein kinase C, as alpha 2-adrenergic stimulation of inositol phosphate production was negligible and the presence of the selective protein kinase C inhibitor RO-31-8220, at concentrations up to 10 microM, had no effect on UK14304-mediated production of phosphatidylbutanol. These results demonstrate that expression of the alpha 2-C10 receptor in a heterologous system can result in receptor regulation of signaling elements that appear not to be primary targets for the receptor in vivo. Such results are important in respect to recent observations that transfection of a single defined receptor into separate cell lines can lead to the regulation of distinct effector systems (Vallar, L., Muca, C., Magni, M., Albert, P., Bunzow, J., Meldolesi, J. and Civelli, O. (1990) J. Biol. Chem. 265, 10320-10326).(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Alpha 2-C10 adrenergic receptors expressed in rat 1 fibroblasts can regulate both adenylylcyclase and phospholipase D-mediated hydrolysis of phosphatidylcholine by interacting with pertussis toxin-sensitive guanine nucleotide-binding proteins. 134 92

The mode of phospholipase C activation in platelet cells induced by didecanoyl (C10)-phosphatidic acid (PA) was investigated with washed rabbit platelets. The C10-PA dose-dependently induced aggregation and serotonin secretion, as well as increases in cytoplasmic free Ca2+ concentration and 1,2-diacylglycerol formation. None of these responses was evoked unless Ca2+ had been added to the platelet suspension. Furthermore, under the conditions of various intracellular Ca2+ concentrations which were set by treatment of the cells with ionomycin and Ca2+, C10-PA promoted 1,2-diacylglycerol formation only at the Ca2+ concentration of 300 nM or higher, whereas thrombin induced the formation even at 100 nM Ca2+. These results suggest that PA activates platelet phospholipase C in cooperation with Ca2+ and contributes to platelet activation through such an effect.
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PMID:Phosphatidic acid with medium-length fatty acyl chains synergistically stimulates phospholipase C with Ca2+ in rabbit platelets. 162 58

To test the hypothesis that agents activating receptors negatively coupled to adenylyl cyclase (AC) can stimulate cell proliferation, we have expressed a human alpha 2-adrenergic receptor (alpha 2-C10) in CCL39 cells and studied the effects of alpha 2-agonists on reinitiation of DNA synthesis in quiescent cells. We report that the alpha 2-agonists epinephrine and clonidine stimulate [3H]-thymidine incorporation in synergy with fibroblast growth factor and that the alpha 2-antagonist yohimbine efficiently inhibits this response. Epinephrine- and clonidine-stimulated DNA synthesis is completely blocked by pertussis toxin and correlates well with the inhibition of prostaglandin E1-stimulated AC. Thus, their action closely resembles the action of serotonin in the same cell system, which is mediated through 5-HT1b receptors. In fact, serotonin- and epinephrine-stimulated DNA synthesis reinitiation is not additive, suggesting that both agents act through a common pathway. Interestingly, alpha 2-agonists also induced a moderate release of inositol phosphates, indicating that alpha 2-adrenergic receptors can interact both with the AC and phospholipase C messenger system. Activation of phosphoinositide (PI) turnover by epinephrine leads to a significant stimulation of Na+/H+ exchange but is insufficient to trigger a mitogenic response in CCL39 cells, as will be discussed. We found no evidence for epinephrine-induced activation of Na+/H+ exchange by a mechanism independent of PI breakdown.Our data show that alpha 2-adrenergic receptors can play a role in the regulation of cell proliferation in an appropriate context; also, the data support the hypothesis that receptors negatively coupled to AC must be taken into account as mediators of growth factor action in fibroblasts, in particular when activated in parallel with receptor tyrosine kinases.
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PMID:Alpha 2-adrenergic agonists stimulate DNA synthesis in Chinese hamster lung fibroblasts transfected with a human alpha 2-adrenergic receptor gene. 198 85

The alpha-adrenergic receptors mediate the effects of epinephrine and norepinephrine on cellular signaling systems via guanine nucleotide binding regulatory proteins (G-proteins). Three alpha-adrenergic receptor subtypes have been cloned: the alpha 1, the alpha 2-C10, and the alpha 2-C4 adrenergic receptors. To investigate functional differences between the different subtypes, we assessed the ability of each to interact with adenylyl cyclase and polyphosphoinositide metabolism by permanently and transiently expressing the DNAs encoding the alpha 1, the alpha 2-C10, and the alpha 2-C4 adrenergic receptors in cells lacking endogenous alpha-adrenergic receptors. Both alpha 2-C10 and alpha 2-C4 couple primarily to inhibition of adenylyl cyclase and to a lesser extent to stimulation of polyphosphoinositide hydrolysis. alpha 2-C10 inhibits adenylyl cyclase more efficiently than alpha 2-C4. Effects of the alpha 2-adrenergic receptors on adenylyl cyclase inhibition and on polyphosphoinositide hydrolysis are both mediated by pertussis toxin-sensitive G-proteins. The major coupling system of the alpha 1-adrenergic receptor is activation of phospholipase C via a pertussis toxin-insensitive G-protein. alpha 1-Adrenergic receptor stimulation can also increase intracellular cAMP by a mechanism that does not involve direct activation of adenylyl cyclase. As with the muscarinic cholinergic receptor family our results show that each of the alpha-adrenergic receptor subtypes can couple to multiple signal transduction pathways and suggest several generalities about the effector coupling mechanisms of G-protein-coupled receptors.
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PMID:Multiple second messenger pathways of alpha-adrenergic receptor subtypes expressed in eukaryotic cells. 215 28

The mechanism of action of the absorption enhancers such as sodium caprate (C10) and decanoylcarnitine (DC) was examined. Both C10 and DC increased the epithelial permeability of fluorescein isothiocyanate dextran 4000 and decreased the transepithelial electrical resistance in Caco-2 cell monolayer. Irrespective of the presence or absence of mucosal calcium, C10 rapidly increased intracellular calcium levels dose-dependently. Compound 48/80, a phospholipase C inhibitor, prevented the increases of the intracellular calcium level and permeability of fluorescein isothiocyanate dextran 4000 by C10. Furthermore, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride, a strong calmodulin inhibitor, also significantly decreased the enhancing effect of C10. These results suggest that C10 releases calcium from intracellular stores via activation of phospholipase C in plasma membrane. The increase of the calcium levels was considered to induce the contraction of calmodulin-dependent actin microfilament, followed by dilatation of the paracellular route. Although DC also increased intracellular calcium levels, neither compound 48/80 nor N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride decreased the enhancing effect of DC. The enhancing mechanisms were different for C10 and DC.
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PMID:Absorption-enhancing mechanism of sodium caprate and decanoylcarnitine in Caco-2 cells. 785 88

The biochemical and functional properties of T. cruzi GP50/55, a novel glycosylphosphatidylinositol (GPI)-anchored membrane antigen have been investigated. A 50-52-kDa thiol proteinase activity could be immunoprecipitated with monoclonal antibodies (mAb) directed against GP50/55 (mAb C10), different from the one reactive with mAbs against lysosomal cysteine proteinase GP57/51. Furthermore, the mAb C10-reactive proteinase corresponded to the GPI-anchored surface antigen since the proteolytic and antigenic activity partitioned to the aqueous phase after Triton X114 phase separation of phosphatidylinositol specific phospholipase C (PI-PLC)-treated parasites. Of several proteins immunoprecipitated by a polyclonal anti-lysosomal cysteine proteinase, an mAb to GP57/51 recognized a 60-kDa protein, whereas mAb C10 recognized antigens ranging between 52 and 50 kDa. The GP50/55 antigen detected by mAb C10 is expressed on the parasite surface whereas the GP57/51 antigen is mainly intracellular. The internal peptide sequence obtained from purified GP50/55 showed that it is more homologous to the prototype of the cysteine proteinases superfamily, papain, than to the two T. cruzi lysosomal cysteine proteinases so far described. Our data indicate that the T. cruzi GP50/55 is a novel GPI-anchored cysteine proteinase and may represent another isoform of this heterogeneous group of proteinases.
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PMID:Trypanosoma cruzi: identification of a membrane cysteine proteinase linked through a GPI anchor. 808 Dec 61

Earlier experiments with animal and human arteries have shown that farnesol, a natural 15-carbon (C15) isoprenoid, is an inhibitor of vasoconstriction (Roullet, J.-B., Xue, H., Chapman, J., McDougal, P., Roullet, C. M., and McCarron, D. A. (1996) J. Clin. Invest. 97, 2384-2390). We report here that farnesol reduced KCl- and norepinephrine-dependent cytosolic Ca2+ transients in fura-2-loaded intact arteries. An effect on Ca2+ signaling was also observed in cultured aortic smooth muscle cells (A10 cells). In these cells, farnesol reduced KCl-induced [Ca2+]i transients and mimicked the inhibitory effect of Ca2+-free medium on the [Ca2+]i response to both 12,13-phorbol myristate acetate, a protein kinase C activator, and thapsigargin, a specific endoplasmic reticulum ATPase inhibitor. Perforated patch-clamp experiments further showed in two vascular smooth muscle cell lines (A10 and A7r5), a reversible, dose-dependent inhibitory effect of farnesol on L-type Ca2+ currents (IC50 = 2.2 microM). Shorter (C10, geraniol) and longer (C20, geranylgeraniol) isoprenols were inactive. L-type Ca2+ channel blockade also occurred under tight (gigaohm) seal configuration using cell-attached, single-channel analysis, thus suggesting a possible action of farnesol from within the intracellular space. We finally demonstrated that farnesol did not affect Ca2+-sensitive pathways implicated in smooth muscle contraction, as tested with alpha-toxin permeabilized arteries. Altogether, our results indicate that farnesol is an inhibitor of vascular smooth muscle Ca2+ signaling with plasma membrane Ca2+ channel blocker properties. The data have implications for the endogenous and pharmacological regulation of vascular tone by farnesol or farnesol analogues.
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PMID:Farnesol inhibits L-type Ca2+ channels in vascular smooth muscle cells. 940 27

Medium chain fatty acids (MCFAs) are used to enhance the permeability of mucosal tissues to hydrophilic drugs, but their mechanism of action is largely unknown. In this study, the absorption-enhancing effects of the sodium salts of two MCFAs, capric acid (C10) and lauric acid (C12), were studied in monolayers of human intestinal epithelial Caco-2 cells. Both MCFAs induced a rapid increase in epithelial permeability to the hydrophilic marker molecule sodium fluorescein. Inhibition of phospholipase C and inhibition or activation of various kinases and buffering of intracellular calcium indicated that the effects on epithelial permeability were mediated through phospholipase C-dependent inositol triphosphate/diacylglycerol pathways. Surprisingly, the inositol triphosphate and diacylglycerol pathways were found to have opposing effects on paracellular permeability. Exposure to the MCFAs also resulted in a concentration dependent reduction of cellular dehydrogenase activity and ATP levels. C10, but not C12, induced redistribution of the tight junction proteins ZO-1 and occludin. These results indicate that the two MCFAs have partially different and more complex mechanisms than previously recognized, which has important implications for their use in vivo.
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PMID:Absorption enhancement through intracellular regulation of tight junction permeability by medium chain fatty acids in Caco-2 cells. 943 99

Although phospholipase C (PLC) is known to be activated by water-insoluble organic solvents, most activity assays have been designed to work in an aqueous milieu. Here a sensitive method is described for the determination of PLC activity in two-phase systems. The assay is based on the hydrolysis of phosphatidylcholine (PC) in chloroform/buffer. The initial rates of the reaction are determined by densitometric quantification of the product 1,2-diacylglycerol after its separation by high-performance TLC and staining with a CuSO4/H3PO4 or p-methoxybenzaldehyde/H2SO4 reagent. The method is examined for the determination of Vmax and Km values of PCs with varying length acyl chains (C10-C18). The comparison of the kinetic parameters with the Vmax and Km values of the same substrates in the conventional titrimetric assay, using sodium deoxycholate for micellization of PC, demonstrates the high efficiency of PLC in the two-phase emulsion system.
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PMID:Activity of phospholipase C in two-phase systems. 1206 25


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