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)

We measured phospholipase activities in both the microsomal and the cytosolic enriched fractions of rat alveolar macrophages by using exogenous arachidonic acid-labeled phospholipids. The microsomal fractions contain a neutral calcium-independent phospholipase A2 (PLA2) which acts on substrates phosphatidylcholine (PC) and phosphatidylinositol (PI), a calcium-independent PLA2 acting on phosphatidylethanolamine (PE), and a neutral calcium-dependent PI-specific PLC. The cytosolic fractions contain calcium-dependent phospholipase: PLA2 that hydrolyses PC at alkaline pH, and a neutral PI-specific phospholipase C (PLC). The largest release of arachidonic acid from PI occurred with the cytosolic fractions at pH 6 in the presence of calcium. That hydrolysis involved a PLA2, and a PLC followed by the action of a diacyglycerol and 2-monoacylglycerol lipases. The cytosol also contains a calcium-independent PLA2 acting on PE. Our investigation shows that rat alveolar macrophages possess a number of phospholipases, as well as diacylglycerol and 2-monoacylglycerol lipases. The above enzymes could play an essential role in the remodeling of membrane phospholipids in resting cells, and the generation of physiologically active lipids in activated cells.
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PMID:Characterization of several phospholipase activities and diacylglycerol/2-monoacylglycerol lipases in rat alveolar macrophages. 189 89

Differentiation with dibutyryl cyclic AMP (dBcAMP) of the human, premonocytic U937 cell line toward a monocyte/granulocyte-like cell results in the cell acquiring an ability to release arachidonate upon stimulation. In contrast, the calcium ionophore ionomycin was able to stimulate phospholipase C, as measured by inositol 1,4,5-trisphosphate formation, to equal extents in both undifferentiated and dBcAMP-differentiated U937 cells. The role and regulation of cytosolic phospholipase A2 (cPLA2) in the production of arachidonate in these cells when either the chemotactic peptide fMLP or ionomycin are used as stimulus were investigated. The ionomycin- and fMLP-stimulated release of arachidonate were sensitive to the cPLA2 inhibitor arachidonyl trifluoromethylketone (IC50 values of 32 and 18 microM, respectively), but were not inhibited by E-6-(bromomethylene)-tetrahydro-3-(1-naphthalenyl)-2 H-pyran-2-one, a bromoenol lactone inhibitor of the calcium-independent phospholipase A2. These results, coupled with the inhibition of ionomycin-induced arachidonate production by electroporation of differentiated cells to introduce an anti-cPLA2, demonstrate that the cPLA2 is the enzyme responsible for arachidonate release in differentiated cells. SDS-PAGE and immunoblot analysis of differentiated cells showed the cells to contain both phosphorylated and unphosphorylated forms of cPLA2 (ratio of about 2: 3). Surprisingly, undifferentiated cells contain 30% more enzyme than differentiated cells and contain a higher percentage (approximately 75%) of the phosphorylated in the absence of stimulation. The inability of undifferentiated cells to produce arachidonate is not due to insufficient intracellular calcium concentrations since ionomycin induces large (820-940 nM) influxes of intracellular calcium in both differentiated and undifferentiated cells. This demonstrates that phosphorylation of cPLA2 andan influx of intracellular calcium are not sufficient to activate the enzyme to produce arachidonate. Instead, activation of a pertussis toxin-sensitive Gi alpha-type G-protein is required as evidenced by the production of arachidonate in undifferentiated cells stimulated with mastoparan, an activator of Gi alpha subunits, in combination with ionomycin. This activation of a Gi alpha-type G-protein is independent of modulations of adenylyl cyclase activity since cellular cAMP levels were not modulated upon treatment with mastoparan and ionomycin.
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PMID:Phosphorylation and calcium influx are not sufficient for the activation of cytosolic phospholipase A2 in U937 cells: requirement for a Gi alpha-type G-protein. 935 62

Arachidonic acid release is an important regulatory component of uterine contraction and parturition, and previous studies showed that lindane stimulates arachidonic acid release from myometrium. The present study partially characterized the enzyme activity responsible for lindane-induced arachidonic acid release in myometrial cells. Lindane released arachidonic acid from cultured rat myometrial cells in concentration- and time-dependent manners. This release was primarily from phosphatidylcholine and phosphatidylinositol, and was independent of intracellular and extracellular calcium. In cells prelabeled with [3H]arachidonic acid, 85% of radiolabel was recovered as free arachidonate and only 5% was recovered as eicosanoids. Pretreatment with the antioxidants Cu, Zn-superoxide dismutase, alpha-tocopherol or Trolox did not significantly modify lindane-induced arachidonic acid release. Pretreatment of cells with the phosphatidylcholine-specific phospholipase C inhibitor D609, phosphatidylinositol-specific phospholipase C inhibitor ET-18-OCH3, or an interrupter of the phospholipase D pathway (ethanol) did not suppress lindane-induced arachidonic acid release. Although these results are consistent with calcium-independent phospholipase A2 activation by lindane, the calcium-independent phospholipase A2 inhibitor bromoenol lactone failed to inhibit lindane-induced arachidonic acid release in myometrial cells, even though bromoenol lactone effectively blocked arachidonic acid release in neutrophils. These results suggest that myometrial cells express a novel, previously unidentified phospholipase that is arachidonate-specific, calcium-independent, insensitive to bromoenol lactone, insensitive to reactive oxygen species activation, shows substrate preference for phosphatidylcholine and phosphatidylinositol, and is stimulated by lindane. Moreover, the data show that the overwhelming majority of arachidonic acid released remains as arachidonate, but that lindane does not significantly inhibit metabolism of arachidonate to eicosanoids.
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PMID:A calcium-independent phospholipase activity insensitive to bromoenol lactone mediates arachidonic acid release by lindane in rat myometrial cells. 1179 14

Our previous in vitro microperfusion studies established that dopamine inhibits sodium chloride transport in the rat medullary thick ascending limb. The present study was designed to determine the intracellular signaling pathway mediating this response. The dopamine D1 receptor agonist fenoldopam (1 microM) inhibited sodium chloride transport in the thick ascending limb by 42+/-5%. The dopamine D1 receptor antagonist R-(+)-7-Chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine-HCl (SCH-23390) completely blocked this effect of fenoldopam. Suppression of protein kinase A activity using either myristoylated protein kinase inhibitor (PKI) or N-[2-(p-Bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide.2HCl (H-89), as well as suppression of phospholipase C activity using 1-(6-((17 beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione (U-73122), had no effect on fenoldopam-dependent inhibition of transport. In contrast, inhibition of phospholipase A2 activity using E-6-(Bromomethylene)tetrahydro-3-(1-naphthalenyl)-2H-pyran-2-one (HELSS) significantly attenuated the effect of fenoldopam by 74%. The cytochrome P-450 monooxygenase inhibitor 17-octadecynoic acid (17-ODYA) and the protein kinase C inhibitor staurosporine both significantly attenuated the effects of fenoldopam by 67%. Exposure to 20-Hydroxy-(5Z, 8Z, 11Z, 14Z)-eicosatetraenoic acid (20-HETE) inhibited transport by 31+/-5%, and this effect was significantly attenuated by 66% in the presence of staurosporine. We propose a signaling pathway in which dopamine activates a calcium-independent phospholipase A2 in the medullary thick ascending limb. Released arachidonic acid is then metabolized to 20-HETE which subsequently increases protein kinase C activity that acts as a final transport effector.
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PMID:Dopamine D1 receptor-dependent inhibition of NaCl transport in the rat thick ascending limb: mechanism of action. 1289 37

Visceral leishmaniasis (VL) is the second-largest parasitic killer disease after malaria. During VL, the protozoan Leishmania donovani induces prostaglandin E2 (PGE2) generation within host macrophages to aid parasite survival. PGE2 significantly influences leishmanial pathogenesis, as L. donovani proliferation is known to be attenuated in PGE2-inhibited macrophages. Here, we report for the first time that signaling via macrophage Toll-like receptor 2 (TLR2) plays an instrumental role in inducing PGE2 release from L. donovani-infected macrophages. This signaling cascade, mediated via the TLR2-phosphatidylinositol 3-kinase (PI3K)-phospholipase C (PLC) signaling pathway, was found to be indispensable for activation of two major enzymes required for PGE2 generation: cytosolic phospholipase A2 (cPLA2) and cyclooxygenase 2 (Cox2). Inhibition of cPLA2, but not secreted phospholipase A2 (sPLA2) or calcium-independent phospholipase A2 (iPLA2), arrested L. donovani infection. During infection, cPLA2 activity increased >7-fold in a calcium-dependent and extracellular signal-regulated kinase (ERK)-dependent manner, indicating that elevation of intracellular calcium and ERK-mediated phosphorylation was necessary for L. donovani-induced cPLA2 activation. For transcriptional upregulation of cyclooxygenase 2, activation of the calcium-calcineurin-nuclear factor of activated T cells (NFAT) signaling was required in addition to the TLR2-PI3K-PLC pathway. Detailed studies by site-directed mutagenesis of potential NFAT binding sites and chromatin immunoprecipitation (ChIP) analysis revealed that the binding of macrophage NFATc2, at the -73/-77 site on the cox2 promoter, induced L. donovani-driven cox2 transcriptional activation. Collectively, these findings highlight the contribution of TLR2 downstream signaling toward activation of cPLA2 and Cox2 and illustrate how the TLR2-PI3K-PLC pathway acts in a concerted manner with calcium-calcineurin-NFATc2 signaling to modulate PGE2 release from L. donovani-infected macrophages.
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PMID:Leishmania donovani-Induced Prostaglandin E2 Generation Is Critically Dependent on Host Toll-Like Receptor 2-Cytosolic Phospholipase A2 Signaling. 2748 Dec 48