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 role of Ca2+ in phospholipid metabolism and arachidonic acid release was studied in guinea pig neutrophils. The chemotactic peptide formylmethionyl-leucyl-phenyl-alanine (fMLP) activated [32P]Pi incorporation into phosphatidylinositol (PI) and phosphatidic acid (PA) without any effects on the labeling of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylserine (PS). This activation was observed in Ca2+-free medium. Even in the neutrophils severely deprived of Ca2+ with EGTA and Ca2+ ionophore A23187, the stimulated labeling was not inhibited. When [3H]arachidonic acid-labeled neutrophils were stimulated by fMLP, a loss of [3H]arachidonic acid moiety in PI and the resultant increase in [3H]arachidonyl-diacylglycerol (DG), -PA, and free [3H]arachidonic acid was marked within 3 min. With further incubation, a loss of [3H]arachidonic acid in PC and PE became significant. These results suggest the activation of phospholipase C preceded the activation of phospholipase A2. In Ca2+-free medium, the decrease in [3H]arachidonyl-PI and the increase in [3H]arachidonyl-PA were only partially inhibited, although the release of [3H]arachidonic acid and a loss of [3H]arachidonyl-PC and -PE was completely blocked. These results show that PI-specific phospholipase C was not as sensitive to Ca2+ deprivation as arachidonic acid cleaving enzymes, phospholipase A2, and diacylglycerol lipase. Ca2+ ionophore A23187, which is known as an inducer of secretion, also stimulated [32P]Pi incorporation into PI and PA, although the incorporation into other phospholipids, such as PC and PE, was inhibited. This stimulated incorporation seemed to be caused by the activation of de novo synthesis of these lipids, because the incorporation of [3H]glycerol into PA and PI was also markedly stimulated by Ca2+ ionophore. But the chemotactic peptide did not increase the incorporation of [3H]glycerol into any glycerolipids including PI and PA. Thus, it is clear that fMLP mainly activates the pathway, PI leads to DG leads to PA, whereas Ca2+ ionophore activates the de novo synthesis of acidic phospholipids. When [3H]arachidonic acid-labeled neutrophils were treated with Ca2+ ionophore, the enhanced release of arachidonic acid and the accumulation of [3H]arachidonyl-DG, -PA with a concomitant decrease in [3H]arachidonyl-PC, -PE, and -PI were observed. Furthermore, the Ca2+ ionophore stimulated the formation of lysophospholipids, such as LPC, LPE, LPI, and LPA nonspecifically. These data suggest that Ca2+ ionophore releases arachidonic acid, unlike fMLP, directly from PC, PE, and PI, mainly by phospholipase A2. When neutrophils were stimulated by fMLP, the formation of LPC and LPE was observed by incubation for more than 3 min. Because a loss of arachidonic acid from PI occurred rapidly in response to fMLP, it seems likely the activation of PI-specific phospholipase C occurred first and was followed by the activation of phospholipase A2 when neutrophils are activated by fMLP...
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PMID:Role of Ca2+ in phosphatidylinositol response and arachidonic acid release in formylated tripeptide- or Ca2+ ionophore A23187-stimulated guinea pig neutrophils. 640 97

Lysophosphatidic acid (LPA; 1-acyl-sn-glycero-3-phosphate) is a platelet-derived lipid mediator that activates its own G-protein-coupled receptor to trigger phospholipase C-mediated Ca2+ mobilization and other effector pathways in numerous cell types. In this study we have examined the structural features of LPA that are important for activation of the Ca(2+)-mobilizing receptor in human A431 carcinoma cells, which show an EC50 for oleoyl-LPA as low as 0.2 nM. When the acyl chain at the sn-1 position is altered, the rank order of potency is oleoyl-LPA > arachidonoyl-LPA > linolenoyl-LPA > linoleoyl-LPA > stearoyl-LPA = palmitoyl-LPA > myristoyl-LPA. The shorter-chain species, lauroyl- and decanoyl-LPA, show little or no activity. Ether-linked LPA (1-O-hexadecyl-sn-glycero-3-phosphate) is somewhat less potent than the corresponding ester-linked LPA; its stereoisomer is about equally active. Deletion of the glycerol backbone causes a 1000-fold decrease in potency. Replacement of the phosphate group in palmitoyl-LPA by a hydrogen- or methyl-phosphonate moiety results in complete loss of activity. A phosphonate analogue with a methylene group replacing the oxygen at sn-3 has strongly decreased activity. All three phosphonate analogues induce cell lysis at doses > 15 microM. Similarly, the methyl and ethyl esters of palmitoyl-LPA are virtually inactive and become cytotoxic at micromolar doses. None of the LPA analogues tested has antagonist activity. Sphingosine 1-phosphate, a putative messenger with some structural similarities to LPA, elicits a transient rise in intracellular [Ca2+] only at micromolar doses; however, cross-desensitization experiments indicate that sphingosine 1-phosphate does not act through the LPA receptor. The results indicate that, although many features of the LPA structure are important for optimal activity, the phosphate group is most critical, suggesting that this moiety is directly involved in receptor activation.
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PMID:Lysophosphatidic acid-induced Ca2+ mobilization in human A431 cells: structure-activity analysis. 773 3

Isolated hepatocytes from fed and starved rats, permeabilized with Staphylococcus aureus alpha-toxin, were incubated with increasing concentrations of radiolabelled fatty acids, in the presence of a saturating concentration of 3-GP. Incorporation of label into LPA, PA and DAG was lower in cells from starved rats than in cells from fed rats, apparently reflecting the lower activity of GPAT after starvation. This enzyme approached saturation at high fatty acid levels and determined the overall flux through the esterification pathway. TAG synthesis, however, was the same in both nutritional states and could not be saturated with fatty acid under the given conditions. Taken together with the observed accumulation of DAG, these data suggest that the rate of TAG synthesis is controlled by the fatty acid supply and, more particularly, by the affinity of DGAT for acyl-CoA.
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PMID:Regulation of triacylglycerol synthesis in permeabilized rat hepatocytes. Role of fatty acid concentration and diacylglycerol acyltransferase. 816 26

Swiss 3T3 fibroblasts were treated with the microtubule-disrupting agent colchicine to study any interaction between microtubule dynamics and actin polymerization. Colchicine increased the amount of filamentous actin (F-actin), in a dose- and time-dependent manner with a significant increase at 1 h by about 130% over control level. Confocal microscopic observation showed that colchicine increased F-actin contents by stress fiber formation without inducing membrane ruffling. Colchicine did not activate phospholipase C and phospholipase D, whereas lysophosphatidic acid did, indicating that colchicine may have a different mechanism of actin polymerization regulation from LPA. A variety of microtubule-disrupting agents stimulated actin polymerization in Swiss 3T3 and Rat-2 fibroblasts as did colchicine, but the microtubule-stabilizing agent taxol inhibited actin polymerization induced by the above microtubule-disrupting agents. In addition, colchicine-induced actin polymerization was blocked by two protein phosphatase inhibitors, okadaic acid and calyculin A. These results suggest that microtubule depolymerization activates stress fiber formation by serine/threonine dephosphorylation in fibroblasts.
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PMID:Colchicine activates actin polymerization by microtubule depolymerization. 926 34

Lysophosphatidate (LPA; 1-acyl-sn-glycero-3-phosphate) is a novel lipid mediator with diverse biological activity. The intracellular mechanisms that mediate the actions of LPA include activation of phospholipase C and protein kinase C (PKC), increases in intracellular Ca2+, inhibition of adenylyl cyclase, and activation of phospholipase D (PLD). We have shown that thyrotropin (TSH) mediated PLD activation involves both the cyclic adenosine monophosphate (cAMP) and PKC pathways. We determined the effects of LPA (10 or 50 microM; 30 minutes) on TSH- and forskolin-mediated cAMP production in FRTL-5 thyroid cells. Basal cAMP was unaffected by LPA. However, both 10 microM and 50 microM LPA inhibited TSH-mediated cAMP production by 66% and 64%, respectively (p < 0.01, ANOVA). A similar inhibition of forskolin-mediated cAMP production was observed following LPA (p < 0.01, ANOVA). After 30-minutes exposure to 50 microM LPA, TSH-mediated iodide uptake (IU) was unaffected. However, 50 microM LPA enhanced TSH-IU after 24-hour exposure by 23%+/-8% (p < 0.03, ANOVA) and inhibited TSH-IU following 72-hour exposure by 43%+/-10% (p < 0.02, ANOVA). There was no effect of LPA on basal IU. To determine whether PLD activation mediated the effects of LPA, PLD activity was examined in FRTL-5 thyroid cells 30 minutes after LPA exposure. While PLD was increased 3.5-fold compared to control values following 50 microM LPA (p < 0.05, ANOVA), no increase in PLD activation was seen following treatment with 10 microM LPA. Preliminary evidence revealed no effect of a protein kinase C inhibitor on LPA inhibition of cAMP generation. To examine the products of PLD activation, we measured the production of phosphatidate (PA) and diacylglycerol (DAG) in FRTL-5 thyroid cells following treatment with 50 microM LPA or 100 microU/mL TSH. Within 1 minute following LPA, a rapid spike of DAG production was observed (1.5- +/- 0.2-fold above basal, p < 0.05, ANOVA). No similar increases in PA or bisPA were demonstrated. However, TSH caused a steady increase in PA and DAG that reached a maximum after 30 minutes. In summary, the effects of LPA on differentiated thyroid function in FRTL-5 thyroid cells are complex. LPA inhibits TSH- and forskolin-mediated cAMP generation most likely via a direct inhibition of adenylyl cyclase, whereas its effects on TSH-IU involve other mechanisms, possibly including PLD activation.
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PMID:The effects of lysophosphatidate on thyrotropin-mediated differentiated thyroid function in FRTL-5 thyroid cells. 1041 Nov 26

PH-20 is a glycoprotein located on the surface of the sperm plasma membrane and on the inner acrosomal membrane. The best understood function of sperm surface PH-20 is its hyaluronidase activity, which results in hydrolysis of the hyaluronic acid-rich cumulus matrix during sperm penetration of this extracellular oocyte investment. In this study, we investigated whether alterations in the secondary and tertiary structures of sperm surface PH-20 would affect its enzyme activity. Proteins were isolated from the sperm plasma membrane by treatment of living cells with phosphatidylinositol-specific phospholipase C (PI-PLC). PH-20 was purified from the PI-PLC released proteins by immunoaffinity chromatography. Two-dimensional electrophoresis of purified PH-20 revealed 6 isoforms with isoelectric points ranging from 5.1 to 6.0. Removal of the N-linked glycans from PH-20 with N-glycosidase F shifted the molecular weight from 64 kd to approximately 54 kd, its deduced molecular weight based on sequence analysis, suggesting that most if not all, of the potential N-glycosylation sites are linked to oligosaccharides. The lectins Con A and PSA recognized purified sperm surface PH-20 after Western blotting, suggesting that mannose is a major sugar within or at the terminal end of the linked glycan. The lectins UEA and LPA did not recognize PH-20 Western blot, suggesting that fucose and sialic acid are not terminal sugars of sperm surface PH-20. Deglycosylation of sperm surface PH-20 resulted in a complete loss of its hyaluronidase activity. The reduction of disulfide bonds with beta-mercaptoethanol or dithiothreitol also resulted in loss of enzyme activity. We conclude that the hyaluronidase activity of sperm surface PH-20 is dependent on structural features established by sulfhydryl linkages, as well as glycosylation.
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PMID:Importance of glycosylation and disulfide bonds in hyaluronidase activity of macaque sperm surface PH-20. 1186 14

Lysophosphatidic acid (LPA; 1-acyl-sn-glycerol-3-phosphate), an abundant constituent of serum, mediates multiple biological responses via G protein-coupled serpentine receptors. Schwann cells express the LPA receptors (Edg receptors), which, once activated, have the potential to signal through G(alphai) to activate p21(ras) and phosphatidylinositol 3-kinase, through G(alphaq) to activate phospholipase C, or through G(q12/13) to activate the Rho pathway. We found that the addition of serum or LPA to serum-starved Schwann cells rapidly (10 min) induced the appearance of actin stress fibers via a Rho-mediated pathway. Furthermore, LPA was able to rescue Schwann cells from apoptosis in a G(alphai)/phosphatidylinositol 3-kinase/MEK/MAPK-dependent manner. In addition, LPA increased the expression of myelin protein P(0) in Schwann cells in a Galpha(i)-independent manner but dependent on protein kinase C. By means of pharmacological and overexpression approaches, we found that the novel isozyme protein kinase Cdelta was required for myelin P(0) expression. Thus, the multiple effects of LPA in Schwann cells (actin reorganization, survival, and myelin gene expression) appear to be mediated through the different G protein-dependent pathways activated by the LPA receptor.
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PMID:Lysophosphatidic acid promotes survival and differentiation of rat Schwann cells. 1252 51

The signaling pathways linking to lysophosphatidic acid-promoted meiotic maturation in mice were studied. When mouse oocyte-cumulus cells complexes were cultured with 10(-5) M lysophosphatidic acid (the LPA group), the rate of oocyte nuclear maturation was significantly increased. Additions of pertussis toxin, genistein, U73122, Ro320432, PD98059 or SB203580 significantly suppressed the increase in lysophosphatidic acid-stimulated nuclear maturation rate. These results suggested that Gi/o-coupled lysophosphatidic acid receptors activate phosphatidylinositol-specific phospholipase C, and result in ERK and MAP kinase activation, which is triggered by diacylglycerol-dependent protein kinase C. When intracellular cAMP concentrations of oocytes in the LPA and control groups were measured using the acetylation assay, the intracellular cAMP concentration of an oocyte in the LPA group was significantly lower than the control oocyte (0.117+/-0.04 fmol/oocyte vs. 0.176+/-0.036 fmol/oocyte, p<0.05). In conclusion, our results suggested that lysophosphatidic acid stimulates phospholipase C through a Gi-protein linked receptor on the surface of mouse cumulus cells and stimulates both extracellular signal-regulated kinase and p38 mitogen-activated kinase, resulting in the closure or loose of gap junctions between cumulus cells and the oocyte. The resultant early decrease of oocyte cAMP levels may promote nuclear maturation of mouse oocytes in vitro.
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PMID:The signaling pathways linking to lysophosphatidic acid-promoted meiotic maturation in mice. 1649 84

In this study, we observed that lysophosphatidylglycerol (LPG) stimulated intracellular calcium ([Ca(2+)](i)) increase in OVCAR-3 human ovarian cancer cells. LPG-stimulated [Ca(2+)](i) increase was inhibited by U-73122 but not by U-73343, suggesting that LPG stimulates calcium signaling via phospholipase C activation. Moreover, pertussis toxin (PTX) almost completely inhibited [Ca(2+)](i) increase by LPG, indicating the activation of PTX-sensitive G-proteins. LPG-induced [Ca(2+)](i) increase was only observed in OVCAR-3 ovarian cancer cells and SK-OV3 ovarian cancer cells among tested several cell types. LPG also induced extracellular signal-regulated kinase (ERK) and Akt phosphorylation in OVCAR-3 ovarian cancer cells. Pertussis toxin did not affect the LPG-induced activation of ERK and Akt phosphorylation. We also found that LPG failed to stimulate NF-kappaB-driven luciferase activity in exogenously LPA(1), LPA(2), or LPA(3)-transfected HepG2 cells. Taken together we suggest that LPG stimulates a membrane bound receptor which is different from well-known LPA receptors (LPA(1), LPA(2), and LPA(3)), resulting in at least two different signaling cascades; one involves a pertussis toxin-sensitive and phospholipase C-dependent [Ca(2+)](i) increase, and the other involves a pertussis toxin-insensitive activation of ERK and Akt in ovarian cancer cells.
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PMID:Effect of lysophosphatidylglycerol on several signaling molecules in OVCAR-3 human ovarian cancer cells: involvement of pertussis toxin-sensitive G-protein coupled receptor. 1716 26

We investigated whether lysophosphatidylethanolamine (LPE) modulates cellular signaling in different cell types. SK-OV3 ovarian cancer cells and OVCAR-3 ovarian cancer cells were responsive to LPE. LPE-stimulated intracellular calcium concentration ([Ca(2+)](i)) increase was inhibited by U-73122, suggesting that LPE stimulates calcium signaling via phospholipase C activation. Moreover, pertussis toxin (PTX) almost completely inhibited [Ca(2+)](i) increase by LPE, indicating the involvement of PTX-sensitive G-proteins. Furthermore, we found that LPE stimulated chemotactic migration and cellular invasion in SK-OV3 ovarian cancer cells. We examined the role of lysophosphatidic acid receptors on LPE-stimulated cellular responses using HepG2 cells transfected with different LPA receptors, and found that LPE failed to stimulate nuclear factor kappa B-driven luciferase. We suggest that LPE stimulates a membrane bound receptor, different from well known LPA receptors, resulting in chemotactic migration and cellular invasion in SK-OV3 ovarian cancer cells.
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PMID:Lysophosphatidylethanolamine stimulates chemotactic migration and cellular invasion in SK-OV3 human ovarian cancer cells: involvement of pertussis toxin-sensitive G-protein coupled receptor. 1771 84


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