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)

Calcium has been implicated as an important factor in prostaglandin production. Phospholipase A2, the enzyme believed to be rate limiting for prostaglandin synthesis, is stimulated by Ca2+; however, the levels of Ca2+ necessary to stimulate phospholipase A2 in cell-free systems are higher than levels achieved in intact cells in response to agonists that stimulate prostaglandin synthesis. We examined the calcium dependency of prostaglandin E2 (PGE2) synthesis in the glomerular mesangial cell. Vasopressin enhanced PGE2 synthesis by mechanisms independent of extracellular Ca2+ concentration. The Ca2+ concentration dependency of PGE2 production was established by rendering cells permeable with digitonin and clamping Ca2+ concentration at various levels. When cytosolic free Ca2+ concentration ([Ca2+]f) was set at levels equal to those measured after stimulation with vasopressin in the intact cell, the PGE2 production by the Ca2+-clamped permeabilized cells was approximately one-half of that obtained in nonpermeabilized cells stimulated with vasopressin. Since stimulation of mesangial cells with vasopressin increases protein kinase C activation as well as [Ca2+]f the effects on PGE2 production of protein kinase C activation with phorbol myristate acetate (PMA) were examined. When permeabilized cells were exposed to Ca2+ concentrations in the range of [Ca2+]f measured in cells treated with vasopressin the addition of PMA approximately doubled PGE2 production. No increase in PGE2 production was observed with PMA when Ca2+ concentration was fixed at basal levels of less than 100 nM. Ca2+-dependent acylhydrolase activity and PGE2 production were inhibited by calmodulin inhibitors, W-7 and compound 48/80. Thus, vasopressin-induced PGE2 production could be explained by a synergistic effect of protein kinase C activation together with an increase in [Ca2+]f. A synergistic action of Ca2+ and PMA on acylhydrolase activity could also be observed in nonpermeabilized cells where A23187 was used to increase [Ca2+]f. The effect of PMA was mimicked by another stimulant of protein kinase C, 1-oleoyl 2-acetylglycerol, albeit with lower potency. Neither PMA nor 1-oleoyl 2-acetylglycerol alone had any effect on acylhydrolase activity. Vasopressin, in the presence of GTP gamma S, stimulated phospholipase C in permeabilized cells when [Ca2+]f was fixed at less than 100 nM, without an associated increase in acylhydrolase activity. This evidence, together with inhibition of acylhydrolase activity with phospholipase A2 inhibitors, dibucaine and mepacrine, indicates that the primary acylhydrolase activity was due to phospholipase A2. The enhanced phospholipase A2 activity observed with protein kinase C activation when [Ca2+]f is increased may be related to phosphorylation of phospholipase A2 itself or phospholipase A2 modulatory proteins. These experiments demonstrate that both Ca2+ and protein kinase C play important roles in the regulation of phospholipase A2 and PGE2 synthesis.
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PMID:Calcium dependency of prostaglandin E2 production in rat glomerular mesangial cells. Evidence that protein kinase C modulates the Ca2+-dependent activation of phospholipase A2. 316 26

1. Rat isolated tracheal smooth muscle preparations respond to phospholipase A2 (PLA2) and phospholipase C (PLC) with contractile responses of highly variable magnitudes. Rat tracheae exposed to PLA2 or PLC for a period of 10-30 min, exhibit airway hyperreactivity (AH) to cooling (10 degrees C), i.e., respond with strong contractile responses. Phospholipase D neither contracted rat tracheae nor induced AH to cooling. 2. PLA2-induced AH to cooling was dependent on the presence of extracellular Ca2+ in the physiological solution. 3. Verapamil, azelastine, diltiazem and TMB-8 (each 10 microM) significantly attenuated PLA2-induced AH. This effect was not shared by nifedipine (10 microM). 4. Bepridil (10 microM), a Ca2+ and calmodulin antagonist, also significantly attenuated AH induced by PLA2. 5. Indomethacin (a cyclo-oxygenase inhibitor), AA-861 (a selective 5-lipoxygenase inhibitor), FPL 55712 (a leukotriene receptor antagonist), methysergide (a 5-hydroxytryptamine D-receptor antagonist) and pyrilamine (a histamine H1-receptor antagonist) exerted little or no effect on PLA2-induced AH to cooling. 6. Atropine significantly attenuated PLA2-induced AH suggesting the participation of acetylcholine. 7. Nordihydroguaiaretic acid (an antioxidant; 5-lipoxygenase inhibitor) and BW 755C (an antioxidant; a dual inhibitor of cyclo-oxygenase and 5-lipoxygenase) significantly attenuated PLA2-induced AH to cooling. 8. In conclusion, these data show that PLA2 (an enzyme involved in the synthesis of Paf-acether, prostaglandins, thromboxanes, leukotrienes, diacylglycerol, superoxide free radicals and lipid peroxides, etc.) induces AH to cooling and acetylcholine in rat trachea. The induction of AH to cooling is dependent on the presence of extracellular Ca2+ and is significantly attenuated by verapamil, diltiazem, bepridil, atropine and azelastine (an antiallergic/antiasthmatic drug).
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PMID:Phospholipase A2 induced airway hyperreactivity to cooling and acetylcholine in rat trachea: pharmacological modulation. 320 72

Several alterations in membrane transport systems are observed in rat and human hypertension. Na+ flux changes are numerous, and cellular homeostasis to Na+ loading is impaired. Transmembrane Ca2+ movements are also numerous but clearly defined by a reduction in Ca2+ binders, a hypersensitivity of membrane phospholipase C, possible increased Ca2+ leak and reduced sensitivity of the Ca2+-pump to calmodulin. The resulting Ca2+ increase within arterial cells can be responsible for increased contractility and tone, leading to hypertension. These functional alterations in membrane transport can be secondary to a few well-defined membrane defects of genetic origin or to a diffuse structural perturbation in membranes involving lipid changes.
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PMID:Cell membrane in hypertension. 328 57

We studied PGE2-release from isolated human gastric mucosal cells. Mucosa was obtained at surgery and cells were dispersed by collagenase and pronase. Centrifugation with Percoll yielded a fraction of light density cells (70-75% parietal cells; 2-4% mast cells) revealing maximal rates of PGE2-release. A radioimmunoassay was used to measure PGE2-release into the incubation medium. Calcium ionophore A23187 which aids calcium transport across membranes caused a 3.5-fold increase of PGE2-release; this effect was abolished in calcium-free incubation medium. PGE2-release was also stimulated by phospholipase C (100 mU/ml) which is known to induce phosphoinositol breakdown, as well as by 1-oleyl-2-acetyl-sn-glycerol (OAG; 10 microM) and by 12-O-tetradecanoyl-13-acetate (TPA; 10 microM) which cause direct activation of protein kinase C without preceding induction of phosphoinositol breakdown. The response to TPA was potentiated by A23187. The calmodulin antagonist naphthalene sulfonamide W 7 reduced PGE2-release in response to A23187 and TPA (IC50: 1 microM). Our data indicate that PGE2-release of human gastric mucosal cells is stimulated by calcium influx as well as by indirect (phospholipase C) and direct (OAG, TPA) activation of protein kinase C. Stimulation of PGE2-release involves calmodulin-mediated mechanisms.
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PMID:[Calcium, phospholipase C and protein kinase C stimulate prostaglandin secretion of isolated gastric mucosa cells of the human]. 347 5

Compound 48/80 inhibited phosphatidylinositol-specific phospholipase C activity from human platelets. Whereas 1 microgram/ml of compound 48/80 slightly stimulated Ca2+-dependent phospholipase A2, higher concentrations led to dose-dependent inhibition of this platelet enzyme. This biphasic effect was confirmed with phospholipases A2 purified from rat liver and human synovial fluid. The aggregation of human platelets induced by ADP and PAF-acether was inhibited by compound 48/80, whereas the aggregation induced by ionophore A23187 was not modified by this compound. These results demonstrate that the inhibition of platelet aggregation by compound 48/80 is not due solely to effects on calmodulin as previously reported, but that inhibition of phospholipases and probably arachidonate mobilization may also be involved.
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PMID:Compound 48/80 is a potent inhibitor of phospholipase C and a dual modulator of phospholipase A2 from human platelet. 360 84

Studies in erythrocytes indicate that staphylococcal alpha-toxin generates discrete transmembrane channels with an effective diameter of 2-3 nm. In cultured, confluent, pig pulmonary arterial endothelial cells we studied the triggering of the arachidonic acid cascade and its dependence on calcium influx, possibly through toxin-created pores. In endothelial cells alpha-toxin time dependently (5-30 min) and dose dependently (0.1-8 micrograms/ml) stimulated the release of radiolabeled arachidonic acid and prostacyclin (PGI2) production in similar amounts as the calcium ionophore A23187 (10 microM). Preincubation of alpha-toxin with neutralizing antibodies abolished the effect. The toxin response was strictly dose dependent on extracellular calcium but not on magnesium. The toxin effect was accompanied by an up to 10-fold increased passive permeability of pulmonary arterial endothelial cells for 45Ca. Interference with calcium-calmodulin function (trifluoperazine, W7) dose dependently reduced production of PGI2, but blockers of physiological calcium channels (verapamil, nimodipine, nisoldipine, and diltiazem) did not. In contrast to the effect of the ionophore A23187, the toxin effect was accompanied by a release of potassium, but in neither system was there a release of lactate dehydrogenase. In addition, alpha-toxin but not ionophore-exposed endothelial cells showed an increased passive influx of small radiolabeled markers (45Ca and [3H]sucrose) but not of large markers [( 3H]inulin and [3H]dextran). These data are consistent with the concept that alpha-toxin triggers the arachidonic acid cascade in pulmonary arterial endothelial cells by calcium influx and suggest that this calcium influx may proceed through toxin-created transmembrane channels.
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PMID:Staphylococcal alpha-toxin-induced PGI2 production in endothelial cells: role of calcium. 391 12

The phospholipase C-activity in crude extracts of bovine blood platelets is strongly inhibited by the calmodulin-inhibitors fluphenazine and calmidazolium in the mM range, and activated by ATP and ADP, but not by AMP. The activating effect is also shown by the nonhydrolysable ATP- and ADP-analogs alpha,beta- and beta,gamma-methyleneadenosine 5'-triphosphate and alpha,beta-methyleneadenosine 5'-diphosphate, thus indicating that it is an allosteric effect. The stimulation of the phospholipase C-activity by ATP is also detectable in some partially purified fractions of the crude platelet extract, but it is abolished on further purification of the enzyme.
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PMID:Phosphatidylinositol-specific phospholipase C from bovine blood platelets. Inhibition by calmodulin-inhibitors--activation by ATP and ADP. 399 81

Within the short period of 5 years, the availability of a variety of specific radioligands has allowed the resolution of alpha 1- and alpha 2-adrenergic receptor populations in many different tissues and enabled researchers to begin investigations of the mechanisms of regulation and coupling of alpha 1 and alpha 2 receptors to their different cellular effector systems. Binding data have demonstrated that the pharmacological properties of each type of alpha receptor are, in general, similar across tissues and species, although there are some differences in the relative affinities of antagonist drugs. Further attempts to subclassify alpha 1 and alpha 2 receptors may be expected in the future. The historical development of the interpretation of [3H]clonidine binding is of interest in this regard. [3H]Clonidine was proposed to label the "agonist state" of the alpha receptor, and then to label alpha 2 receptors. It is now thought that it labels the agonist state of alpha 2 receptors. Might it actually label a subpopulation of alpha 2 receptors or just the agonist state of that subpopulation? Alpha-1 receptors by and large appear to occur in a single-affinity state with respect to both agonists and antagonists. By comparison, alpha 2 receptors may exist in multiple-affinity states reflecting the ability of the alpha 2 binding site protein to complex to additional membrane proteins which themselves are receptors for the physiological substrates GTP, Na+, Mg2+, and possibly Ca2+-calmodulin. Binding studies have also strongly indicated that alpha 2 receptors in most, if not all, tissues are probably coupled in an inhibitory manner to adenylate cyclase, as has been demonstrated in platelets, adipocytes, and NG 108-15 cells. Clearly the present status of alpha-receptor research has left many questions unresolved. We still have no idea what membrane effector system and associated second messenger is coupled to the alpha 1 receptor. The prevailing belief is that Ca2+ and the membrane Ca2+ channel fulfill these roles. However, others have suggested that phosphoinositide turnover represents the proximal receptor response, and indeed a membrane-bound phospholipase C may play an analogous role to adenylate cyclase for other adrenergic receptors (Putney et al., 1980). There is, however, some evidence that in some situations alpha 1 receptors may directly stimulate adenylate cyclase, and guanine nucleotide modulation of agonist affinities at alpha 1-receptor sites has been reported. The significance of these data and reported modulatory effects of Na+ at alpha 1 receptors (Glossmann and Presek, 1979; Glossmann et al., 1981) is still to be resolved.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Characterization of alpha 1- and alpha 2-adrenergic receptors. 631 99

A calmodulin-Ca2+-stimulated cyclic nucleotide phosphodiesterase (EC 3.1.4.17) which hydrolyzed both cGMP and cAMP has been purified about 2000-fold from ovaries of the amphibian Xenopus laevis. Gel filtration through Sephadex G-200 indicated a molecular weight of 140,000. A single, major protein band of molecular weight 66,000 was observed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In addition to the stimulation by calmodulin-Ca2+, the enzyme was activated 5- to 10-fold by proteolysis and by certain phospholipids. Trypsin activation of the enzyme caused a reduction in the native molecular weight to 90,000 and a loss of the capacity to be stimulated by calmodulin-Ca2+ or by phospholipids. The phosphodiesterase was stimulated by low concentrations (0.1 microgram/ml) of lysophosphatidylcholine and lysophosphatidylethanolamine. This response did not require calcium ions. Phosphatidylinositol, fatty acids, progesterone, and phospholipase C had little or no effect on activity. Simultaneous addition of 1 mM 2-chloro-10-(3-aminopropyl)phenothiazine and lysophosphatidylcholine to the enzyme did not diminish the stimulatory effect of the phospholipid. The activation of the enzyme by all three agents resulted in an increase in the maximum velocity of the reaction without significant modification of the apparent Km values for cGMP (5 microM) or cAMP (30 microM). It was suggested that trypsin removed an inhibitory domain from the enzyme and that calmodulin and phospholipids interact with this same domain, eliminating its capacity to inhibit the active center of the enzyme.
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PMID:Properties of a cyclic nucleotide phosphodiesterase of amphibian oocytes that is activated by calmodulin and calcium, by tryptic proteolysis, and by phospholipids. 632 99

The present study examined (a) the source of arachidonic acid for Ca2+-stimulated renal inner medullary prostaglandin synthesis, (b) the Ca2+-dependence of enzymes of the phospholipase A2 and C pathways, and (c) the role of calmodulin in these Ca2+ actions. Ca2+ plus the ionophore A23187 stimulated (2-4-fold) release of labeled arachidonate, diglyceride, prostaglandin E2 or F2 alpha from inner medullary slices with a concomitant fall in labeled phosphatidylcholine, phosphatidylinositol, and phosphatidylethanolamine. The calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide hydrochloride (W-7) (10-100 microM) abolished or suppressed Ca++-stimulated immunoreactive prostaglandin E, labeled arachidonate and prostaglandin release, and the fall in labeled phospholipids but did not suppress labeled diglyceride or inositol accumulation. Studies in subcellular fractions demonstrated a particulate phospholipase A2 activity and a phosphatidylinositol-specific phospholipase C activity which was predominantly soluble (80%). W-7 or trifluoperazine (25 microM) abolished Ca2+-stimulated phospholipase A2 activity and particulate phospholipase C activity but were without effect on soluble phospholipase C. W-7 (100 microM) was without effect on Ca2+-stimulated diglyceride lipase and phosphatidic acid-specific phospholipase A2 activities. Hypertonic urea at concentrations that pertain in the inner medulla of hydropenic rats in vivo inhibited Ca2+-induced increases in labeled arachidonate release and immunoreactive prostaglandin E in slice incubates and Ca2+-responsive phospholipase C and A2. The results are consistent with the involvement of phospholipase A2, C, or both in the Ca2+ (+A23187)-stimulated release of free arachidonate for prostaglandin synthesis and support a role for calmodulin in Ca2+ activation of phospholipase A2 and particulate phospholipase C.
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PMID:Ca2+.Calmodulin-dependent release of arachidonic acid for renal medullary prostaglandin synthesis. Evidence for involvement of phospholipases A2 and C. 640 36


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