Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. In RAW 264.7 macrophages, lipopolysaccharide (LPS) and gamma-interferon (IFN gamma) alone or in combination stimulated the induction of nitric oxide synthase (iNOS) activity and increased the expression of the 130 kDa isoform of NOS. 2. LPS-induced NOS activity was reduced by incubation with CD14 neutralising antibodies and abolished in macrophages deprived of serum. 3. LPS stimulated a small increase in protein kinase C (PKC) activity in RAW 264.7 macrophages which was dependent on the presence of serum. However, IFN gamma did not potentiate LPS-stimulated PKC activity. 4. The protein kinase C inhibitor, Ro-318220, abolished both LPS- and IFN gamma-stimulated protein kinase C activity and the induction of NOS activity. 5. LPS- and IFN gamma-induced NOS activity was reduced by the tyrosine kinase inhibitor genestein. Genestein also reduced LPS-stimulated protein kinase C activity but did not affect the response to the protein kinase C activator, tetradecanoylphorbol acetate (TPA). 6. Nicotinamide, an inhibitor of poly-ADP ribosylation, abolished LPS- and IFN gamma-induced NOS activity. 7. Brefeldin A, an inhibitor of a factor which stimulates nucleotide exchange activity on the 21 kDa ADP-ribosylation factor, ARF, reduced LPS- and IFN gamma-induced NOS activity by approximately 80%. 8. These results suggest the involvement of protein kinase C, tyrosine kinase and poly-ADP ribosylation pathways in the regulation of the induction of nitric oxide synthase in RAW 264.7 macrophages by LPS and IFN gamma.
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PMID:Protein kinase C and tyrosine kinase pathways regulate lipopolysaccharide-induced nitric oxide synthase activity in RAW 264.7 murine macrophages. 753 21

Phosphatidylcholine-specific phospholipase D (PLD) is an important signalling phospholipase in mammalian cells. Recently, PLD activity has been shown to be positively regulated by the GTP-binding protein ARF (ADP-ribosylating factor). In the present work, we document the presence of a factor negatively regulating PLD activity in bovine brain cytosol. The inhibitory factor is characterized as a large protein or a complex of proteins with a molecular mass higher than 300 kDa. Using permeabilized and pre-permeabilized HL-60 cells depleted of their cytosol, we demonstrate that the inhibitor acts on GTP[S]-stimulated PLD activity. This effect is immediate, persistent and dose dependent for GTP[S]-stimulated PLD. Different possibilities for a mechanism of action of the inhibitory factor on the regulation of GTP binding of ARF were investigated. This inhibitory factor is not the guanine-dissociating inhibitor (GDI) for the small G-binding proteins Rho (Rho-GDI), reported to be a PLD inhibitor, since specific antibodies against this protein did not recognize a protein in the peak containing the inhibitory factor for PLD activity. Furthermore, the inhibitory factor does not prevent the binding of GTP[S] to ARF in the presence of HL-60 membranes. This excludes its possible role as an inhibitor of an ARF/guanine exchange factor. The inhibitory factor not only inhibits a pathway of PLD through GTP[S] activation in particular of the small GTP-binding protein, ARF, but it also inhibits PLD activated via either protein kinase C (PKC) or tyrosine kinase activation. The inhibitory factor also decreases PLC activity and this effect seems to be secondary to the inhibition of PLD activity. We discuss a mechanism of action of the inhibitor on PLD and the importance of this enzyme activity for membrane traffic.
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PMID:A soluble protein negatively regulates phospholipase D activity. Partial purification and characterization. 762 81

Human neutrophil PLD activity stimulated with GTP-gamma-S was reconstituted with recombinant ARF1 in cytosol-depleted cells. PMA-pretreatment of intact cells greatly enhanced the subsequent reconstitution of the ARF1-regulated PLD activity. This enhancement was only observed provided that the intact cells were pretreated with PMA, suggesting the stable recruitment of a cytosolic component, presumably protein kinase C, to the membranes. rARF1-reconstituted PLD activity was not dependent on MgATP, but could be considerably enhanced by MgATP. Maximal effects of MgATP were seen at 1 mM. This enhancement by MgATP could not be attributed to protein kinase C. Neomycin was found to inhibit ARF1-regulated PLD activity suggesting the requirement for polyphosphoinositides. We conclude: (i) that many of the observed effects of PMA may be dependent on the presence of the small GTP-binding protein, ARF, and (ii) polyphosphoinositides are required for ARF1-stimulated PLD activity.
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PMID:ARF1-regulated phospholipase D in human neutrophils is enhanced by PMA and MgATP. 792 57

Neutrophils play a major role host defense against invading microbes. Recent studies have emphasized the importance of the phospholipase D (PLD) in the signalling cascade leading to neutrophil activation. Phospholipase D catalyzes the hydrolysis of phospholipids to generate phosphatidic acid with secondarily generation of diradylglycerol; both of these products have been implicated as second messengers. Herein, we discuss the regulation and the biochemistry of the receptor-regulated PLD in human neutrophils. In vivo and in vitro studies suggest an activation mode in which initial receptor-linked activation of phospholipase C generates diacylglycerol and inositol trisphosphate. The resulting calcium flux along with the diacylglycerol activate a conventional isoform of protein kinase C (PKC), probably PKC beta 1. This PKC, in turn phosphorylates a plasma membrane component resulting in PLD activation and a second outpouring of diradylglycerol. The small GTP-binding proteins, RhoA and ARF, also participate in this process, and synergize with a 50 kDa cytosolic regulatory factor.
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PMID:Biochemistry and cell biology of phospholipase D in human neutrophils. 868 27

Recent evidence in whole cells has implicated ceramide in the regulation of phospholipase D (PLD). In intact HL-60 cells, phorbol myristate acetate (PMA) activated PLD as measured by [3H]palmitate-labeled phosphatidylcholine conversion to phosphatidylethanol in the presence of 2% ethanol. C6-Ceramide completely inhibited PLD activation after 4 h of treatment and was maximally active at 10 microM. The activity was structurally specific in that the structural analogs 4,5-dihydro-C6-ceramide and dioctanoylglycerol were inactive. Although ceramide inhibited PMA-induced activation of PLD, it did not inhibit translocation of protein kinase C (PKC) to the membrane in response to PMA. In a cell-free system, we confirmed that PLD is activated by guanosine 5'-O-(3-thiotriphosphate (GTPgammaS); however, ceramide had no effect on this activity under a variety of conditions. Activation of PLD by GTPgammaS was synergistically enhanced by the addition of PKC activators. This upstream effect was inhibited rapidly and specifically by ceramide (30 microM). Recombinant ARF plus PKCalpha substituted for crude cytosol in the activation of PLD, and this activity was inhibited by C6-ceramide. Taken together, these data show that ceramide interferes with PKC-mediated activation of PLD.
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PMID:Ceramide inhibits phospholipase D in a cell-free system. 879 52

Hydrolysis of phosphatidylcholine via receptor-mediated stimulation of phospholipase D produces phosphatidate that can be converted to lysophosphatidate and diacylglycerol. Diacylglycerol is an activator of protein kinase C, whereas phosphatidate and lysophosphatidate stimulate tyrosine kinases and activate the Ras-Raf-mitogen-activated protein kinase pathway. These three lipids can stimulate cell division. Conversely, activation of sphingomyelinase by agonists (e.g., tumor necrosis factor-alpha) causes ceramide production that inhibits cell division and produces apoptosis. If ceramides are metabolized to sphingosine and sphingosine 1-phosphate, then these lipids can stimulate phospholipase D and are also mitogenic. By contrast, ceramides inhibit the activation of phospholipase D by decreasing its interaction with the G-proteins, ARF and Rho, which are necessary for its activation. In whole cells, ceramides also stimulate the degradation of phosphatidate, lysophosphatidate, ceramide 1-phosphate, and sphingosine 1-phosphate through a multifunctional phosphohydrolase (the Mg(2+)-independent phosphatidate phosphohydrolase), whereas sphingosine inhibits phosphatidate phosphohydrolase. Tumor necrosis factor-alpha causes insulin resistance, which may be partly explained by ceramide production. Cell-permeable ceramides decrease insulin-stimulated glucose uptake in 3T3-L1 adipocytes after 2-24 h, whereas they stimulate basal glucose uptake. These effects do not depend on decreased tyrosine phosphorylation of the insulin receptor and insulin receptor substrate-1 or the interaction of insulin receptor substrate-1 with phosphatidylinositol 3-kinase. They appear to rely on the differential effects of ceramides on the translocation of GLUT1-and GLUT4-containing vesicles. It is concluded that there is a significant interaction and "cross-talk" between the sphingolipid and glycerolipid pathways that modifies signal transduction to control vesicle movement, cell division, and cell death.
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PMID:"Cross talk" between the bioactive glycerolipids and sphingolipids in signal transduction. 896 Mar 53

Serial deletions of the N-terminal 319 amino acids of rPLD1 expressed in COS-7 cells resulted in increased basal PLD activity. Incubation of the cells with phorbol myristate acetate increased the activity of endogenous and wild-type rPLD1. The mutant rPLD1 with deletion of the first 50 amino acids responded to the phorbol ester, however, rPLD1 with deletions of 115 amino acids or more did not. In cells in which constitutively active V14RhoA was co-expressed with the mutant PLDs, stimulation of PLD activity was observed with all deletion mutants. In membranes from COS-7 cells in which the mutant PLDs were expressed, only the mutant with deletion of 50 N-terminal amino acids responded to added protein kinase C-alpha and phorbol ester, in agreement with the in vivo studies. When myristoylated ADP-ribosylation factor 3 (mARF3) was added together with guanosine 5'-3-O-(thio)triphosphate, all mutants showed stimulation of PLD activity. It is concluded that the site of interaction of protein kinase C with rPLD1 is located in the N-terminal region and that Rho and ARF interact at other sites.
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PMID:Definition of the protein kinase C interaction site of phospholipase D. 951 32

The mammalian phosphatidylcholine-specific phospholipase D (PLD) enzymes PLD1 and PLD2 have been proposed to play roles in signal transduction and membrane vesicular trafficking in distinct subcellular compartments. PLD1 is activated in a synergistic manner in vitro by protein kinase C-alpha, ADP-ribosylation factor 1 (ARF1), and Rho family members. In contrast, PLD2 is constitutively active in vitro. We describe here molecular analysis of PLD2. We show that the NH2-terminal 308 amino acids are required for PLD2's characteristic high basal activity. Unexpectedly, PLD2 lacking this region becomes highly responsive to ARF proteins and displays a modest preference for activation by ARF5. Chimeric analysis of PLD1 and PLD2 suggests that the ARF-responsive region is in the PLD carboxyl terminus. We also inserted into PLD2 a region of sequence unique to PLD1 known as the "loop" region, which had been proposed initially to mediate effector stimulation but that subsequently was shown instead to be required in part for the very low basal activity characteristic of PLD1. The insertion decreased PLD2 activity, consistent with the latter finding. Finally, we show that the critical role undertaken by the conserved carboxyl terminus is unlikely to involve promoting PLD association with membrane surfaces.
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PMID:Molecular analysis of mammalian phospholipase D2. 986 70

As phospholipase D (PLD) activation has been associated with mitogenic signalling in several cell types, we tested an association between adrenergic activation of PLD and cellular proliferation in primary cultures of rat cortical astrocytes. In 2-week old cultures, PLD activation by noradrenaline (EC50: 0.49 microM) was inhibited by prazosin, a specific antagonist at alpha1-adrenergic receptors (IC50: 0.23 microM). Adrenergic PLD activation was not affected by genistein, an inhibitor of tyrosine kinases, or by Ro 31-8220, an inhibitor of protein kinase C (PKC), but was dose-dependently depressed in the presence of brefeldin A (1-100 microg/ml), an inhibitor of ARF activation. In experiments measuring cell proliferation, noradrenaline potently (EC50: 20 nM) reduced [3H]thymidine incorporation to 20-30% of basal values. This action was mimicked by the beta-specific agonist isoprenaline and was inhibited by the beta-antagonist propranolol in a concentration-dependent manner. The alpha1-adrenergic agonists, phenylephrine and methoxamine, also reduced DNA synthesis. The adrenergic inhibition of astroglial DNA synthesis was not reduced, but further potentiated in the presence of brefeldin A, ethanol, and 1- and 2-butanol; 1-butanol, a substrate of PLD, was equally effective as 2-butanol, a non-substrate. We conclude that adrenergic PLD activation in astrocytes is not involved in mitogenic signalling. The involvement of ARF in the activation of PLD via alpha1-adrenoceptors indicates a role in protein trafficking.
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PMID:Adrenergic modulation of astroglial phospholipase D activity and cell proliferation. 1035 May 67

The activity on ARF of the guanine nucleotide exchange factor ARNO depends on its membrane recruitment, induced by binding of its PH domain to phosphoinositides. A polycationic C-terminal extension to the PH domain might also contribute to its specific binding to phosphatidylinositol 4,5-bisphosphate [(4,5)PIP2] and to phosphatidylinositol 3,4,5-trisphosphate [(3,4,5)PIP3], and to ionic binding to other acidic lipids. We have analyzed in vitro the relative contributions to phospholipid binding of the PH domain and C-terminal extension by cosedimentation of "PH+C domain" and "nominal PH domain" protein constructs including or not including the polycationic C-terminus, with sucrose-loaded unilamellar vesicles made of equal proportions of the neutral lipids phosphatidylcholine and phosphatidylethanolamine, and supplemented or not with 30% acidic phosphatidylserine (PS) and 2% of various phosphoinositides. Binding was measured as a function of the vesicle concentration and of the medium ionic strength. Both proteins bound with higher affinity to (3,4,5)PIP3 than to (4,5)PIP2, the selectivity for (3,4,5)PIP3 being highest for the nominal PH domain. We observed also a clear selectivity of (3,4,5)PIP3 over (4,5)PIP2 for stimulating the activity of ARNO on ARF with vesicles containing 10% PS and 1% PIP2 or PIP3. Our data suggest that the PH domain provides the specific phosphoinositide binding site and some unspecific ionic interaction with acidic PS, whereas the polybasic C domain contributes to binding mainly by unspecific ionic interactions vith PS. Phosphorylation by protein kinase C of a serine in the C domain reduces the ionic affinity of the PH+C domain for PS, but does not affect the phosphoinositide specificity.
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PMID:Binding of the PH and polybasic C-terminal domains of ARNO to phosphoinositides and to acidic lipids. 1080 41


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