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)

The coagulation protein thrombin has been shown to stimulate multiple endothelial-cell (EC) functions, including production of platelet-derived growth factor and of platelet-activating factor (PAF), and neutrophil adhesion. We have found that thrombin causes increased binding of monocytic cells (U937 cells and normal human monocytes) to cultured EC of various species. Maximum adhesion of monocytes to pig aortic EC occurred 6 h after thrombin treatment and remained elevated through 24 h. Stimulation of adherence by bovine alpha-thrombin was half-maximal at 15 units/ml, and reached a plateau at 50 units/ml. Catalytically inactive thrombin (phenylmethanesulphonyl fluoride-treated) had no effect on monocyte adhesion to EC. Heparin, but not the endotoxin antagonist polymyxin B, suppressed the stimulation of adhesion by thrombin without altering basal adhesion. Two lines of evidence suggested that protein kinase C (PKC) was involved in the intracellular signalling to increase monocyte adhesion to EC. First the PKC activator phorbol 12-myristate 13-acetate (PMA) stimulated monocytic-cell adhesion to EC at a dose consistent with stimulation of PKC (half-maximal response at 1-3 nM) and with a time course similar to that for thrombin stimulation (maximal by 4 h). Diacylglycerol, a physiological activator of PKC, also stimulated U937-cell adhesion to EC. Secondly, H7, a PKC inhibitor, completely blocked stimulation of monocyte adhesion to EC by thrombin or PMA. The structural analogue of H7, HA1004, which preferentially inhibits cyclic-AMP- and cyclic-GMP-dependent protein kinases, had no effect on stimulated monocyte adhesion. The PKC inhibitor also blocked the stimulation of monocyte adhesion to EC by interleukin-1 and endotoxin, but did not alter the basal level of monocyte binding to unstimulated EC. Thrombin stimulation of monocyte adhesion differed from the reported stimulation of neutrophil adhesion by thrombin in that the latter process reached a maximum in minutes rather than hours. In addition, neither PAF itself nor agents known to stimulate PAF production by EC, such as arachidonate and the Ca2+ ionophore A23187, had any effect on monocyte adhesion. These results demonstrate a PKC-dependent cytokine-like action of the coagulation protein thrombin in modulating monocytic-cell adhesion to EC, a phenomenon of potential importance in many pathological and physiological processes.
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PMID:Thrombin causes increased monocytic-cell adhesion to endothelial cells through a protein kinase C-dependent pathway. 251 8

Heparin is a complex glycosaminoglycan that inhibits the proliferation of several cell types in culture and in vivo. To begin to define the mechanism(s) by which heparin exerts its antiproliferative effects, we asked whether heparin interferes with the expression of the growth factor-inducible protooncogenes c-fos and c-myc. We show that heparin suppressed the induction of c-fos and c-myc mRNA by serum in murine (BALB/c) 3T3 fibroblasts. Using purified mitogens, we further show that suppression was most marked when protooncogene expression was induced by phorbol 12-myristate 13-acetate, an activator of protein kinase C. By contrast, there was little or no suppression when the cells were stimulated by epidermal growth factor, which, in these cells, utilizes a protein kinase C-independent pathway for the induction of gene expression. Heparin also inhibited the change in cell morphology induced by the phorbol ester but had no effect on the morphological change induced by epidermal growth factor and agents that raise intracellular cAMP. Heparin did not inhibit intracellular protein kinase C activity, phorbol ester-induced down-regulation of protein kinase C, or phosphorylation of the 80-kDa intracellular protein kinase C substrate. These results suggest that heparin inhibits a protein kinase C-dependent pathway for cell proliferation and suppresses the induction of c-fos and c-myc mRNA at a site distal to activation of the kinase.
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PMID:Heparin suppresses the induction of c-fos and c-myc mRNA in murine fibroblasts by selective inhibition of a protein kinase C-dependent pathway. 254 34

Basic fibroblast growth factor (FGF) is synthesized as a phosphoprotein by both bovine capillary endothelial and human hepatoma cells in culture. Because basic FGF is characterized by its high affinity for heparin and its association in vivo with the extracellular matrix, we examined the possibility that the phosphorylation of this growth factor by purified protein kinase C (PK-C) and the catalytic subunit of cAMP-dependent protein kinase-A (PK-A) can be modulated by components of the extracellular matrix. Heparin and other glycosaminoglycans (GAGs) inhibit the ability of PK-C to phosphorylate basic FGF. In contrast, heparin can directly increase the phosphorylation of basic FGF by PK-A. While fibronectin, laminin, and collagen IV have no effect on the ability of PK-C to phosphorylate basic FGF, they all can inhibit the effects of PK-A. Thus, there is a differential effect of extracellular matrix-derived proteins and GAGs on the phosphorylation of basic FGF. The enhanced phosphorylation of basic FGF that is mediated by heparin is associated with a change in the kinetics of the reaction and the identity of the amino acid targeted by this enzyme. The amino acids that are targeted by PK-C and PK-A have been identified by phosphopeptide analyses as Ser64 and Thr112, respectively. In the presence of heparin, basic FGF is no longer phosphorylated by PK-A at the usual PK-A consensus site (Thr112), but instead is phosphorylated at the canonical PK-C site (Ser64). Accordingly, heparin inhibits the phosphorylation of basic FGF by PK-C presumably by masking the PK-C dependent consensus sequence surrounding Ser64. Thus, when basic FGF is no longer phosphorylated by PK-A in the receptor binding domain (Thr112), it loses the increased receptor binding ability that characterizes PK-A phosphorylated basic FGF. The results presented here demonstrate three novel features of basic FGF. First, they identify a functional effect of the binding of heparin to basic FGF. Second, they establish that the binding of heparin to basic FGF can induce structural changes that alter the substrate specificity of protein kinases. Third, and perhaps most important, the results demonstrate the existence of a novel interaction between basic FGF, fibronectin, and laminin. Although the physiological significance of this phosphorylation is not known, these results clearly suggest that the biological activities of basic FGF are regulated by a complex array of biochemical interactions with the proteins, proteoglycans, and glycosaminoglycans present in the extracellular milieu and the cytoplasm.
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PMID:Differential effects of heparin, fibronectin, and laminin on the phosphorylation of basic fibroblast growth factor by protein kinase C and the catalytic subunit of protein kinase A. 259 18

Protein-kinase activities in rabbit ciliary process tissue were characterized and quantitated using histone, casein, and myosin light chain as substrates. At least four different protein-kinase activities were separated and identified in the supernatant (soluble) and in the particulate fraction using DEAE-cellulose ion-exchange chromatography. Typical activities of the protein kinases in ciliary processes dissected from one eye were as follows: in the supernatant fraction; protein kinase C, 185.0 pmol min-1; cyclic AMP-dependent protein kinase type II, 34.0 pmol min-1; casein kinase type II, 85.1 pmol min-1; protein kinase M, 9.8 pmol min-1: in the particulate fraction; protein kinase C, 55.1 pmol min-1; cyclic AMP-dependent protein kinase type II, 12.5 pmol min-1; casein kinase type II, 13.4 pmol min-1, and protein kinase M, 5.5 pmol min-1. No cyclic GMP-dependent and no calmodulin-dependent protein-kinase activities were detectable using histone, casein or myosin light chain as substrates. The apparent molecular weight of protein kinase C as estimated by exclusion chromatography on a column of Sephadex G-200 was about 90,000. Inhibitory and stimulatory effects of recently synthesized isoquinolinesulfonamide derivatives (H-7 and H-8), heparin, and polylysine were studied in ciliary process protein kinases. H-7 and H-8 were potent inhibitors of cyclic AMP-dependent protein kinase, protein kinase C and protein kinase M, (IC50 less than 10 microM) but had no inhibitory effects on casein kinase. Heparin at 4 micrograms ml-1 inhibited casein kinase activity almost completely without affecting cyclic AMP-dependent or protein kinase C activities. Poly D- or L-lysine were both found to activate (approximately double) casein kinase activity at 40 micrograms ml-1, but did not significantly activate cyclic AMP-dependent protein kinase or protein kinase C. These results provide basic information on the protein kinase enzymes in the ciliary process and show that protein kinase C is the major kinase in this tissue. This suggests a possible role of the Ca2+ and protein kinase C system in transport functions of ciliary processes and in the regulatory mechanism of aqueous-humor formation additional to the already established importance of the cyclic AMP-dependent protein-kinase enzyme.
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PMID:Analysis of protein kinase activities in rabbit ciliary processes: identification and characterization using exogenous substrates. 347 66

We studied the inhibitory effects of heparin, thrombin inhibitor, and protein kinase C (PKC) inhibitor on basal and thrombin-induced preproendothelin-1 (prepro-ET-1) and proto-oncogene c-fos mRNA expression in cultured bovine endothelial cells (ECs). Northern blot analysis using cDNA for bovine prepro-ET-1 as a probe showed that heparin lowered not only the basal but also the stimulated expression of prepro-ET-1 mRNA by thrombin. A selective thrombin inhibitor (argatroban) and a PKC inhibitor (staurosporine) also inhibited thrombin-induced but not basal prepro-ET-1 mRNA expression. Heparin similarly inhibited thrombin-induced c-fos proto-oncogene mRNA expression in ECs. These data suggest that heparin, in addition to its antithrombin effect, has an inhibitory effect on prepro-ET-1 mRNA expression, possibly via a PKC-dependent pathway.
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PMID:Heparin inhibits endothelin-1 and proto-oncogene c-fos gene expression in cultured bovine endothelial cells. 750 97

The effects of some protein kinase effectors on phosphohydrolase and transport activities of yeast vacuoles have been studied. The platelet-activating factor (PAF), a plant vacuolar protein kinase C stimulator, had a protonophoric and membrane-damaging effects on yeast vacuoles and inhibited the ATP-dependent delta mu H+ formation and ATP-dependent secondary transport but stimulated the ATPase and pyrophosphatase hydrolase activities by abrogating proton control. PAF increasing the tonoplast permeability for the corresponding substrates also stimulated pyrophosphatase, polyphosphatase and alkaline phosphatase activities. Lysolipid sphingosine, a plant vacuolar protein kinase C inhibitor, poorly stimulated the ATPase activity and the ATP-dependent formation of Em in isolated yeast vacuoles, while the pyrophosphatase activity increased by 200%. Other hydrolase activities tested were insensitive to the effect of the lysolipid. Sphingosine inhibited the ATP-dependent citrate transport only insignificantly. Heparin, an effective casein kinase inhibitor, suppressed the ATPase and polyphosphatase activities in isolated yeast vacuoles. The polyphosphatase activity was inhibited both in the vacuolar sap and the tonoplast solubilized by a Zwittergent TM-314, in contrast with the ATPase activity which was inhibited by heparin only in isolated vacuoles. Heparin is suggested to inhibit polyphosphatase by directly influencing the enzyme.
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PMID:[The effect of PAF, sphingosine and heparin on certain phosphohydrolase and transport activity of yeast vacuoles]. 794 17

Vascular smooth muscle cells (SMC) do not represent a homogeneous population (Schwartz et al., 1990, Am. J. Pathol. 136: 1417-1428). Cellular clones resistant to the antiproliferative activity of heparin were isolated from rat aortic SMC cultures (Pukac et al., 1990, Cell Regul., 1:435-443; San Antonio et al., 1993, Arterioscler. Thromb., 13:748-757) and from explant of human arterial restenotic lesions (Chan et al., 1993, Lancet, 341:341-342). We have shown in the present study that long-term treatment (growth medium supplemented with 200 micrograms/ml heparin, from the second to the tenth passage) of rat aortic SMC, without cell cloning, resulted in a significant loss of sensitivity to the growth inhibition by heparin and its derivatives. The heparin resistance was stable after growing cells for two passages in heparin-free medium, suggesting the selection of a particular phenotype. We tried to characterize these cells and to determine the causes of the resistance to the growth inhibition by heparin. Heparin-treated SMC (HT-SMC) were smaller than their control culture at the same passage, expressed less alpha-SM actin, and did not overgrow after reaching confluence. As in the heparin-resistant clones (San Antonio et al., 1993, Cell Regul., 1:435-443) expression of alpha-SM actin could be increased in HT-SMC by heparin addition before Western blotting. Heparin resistance was associated with a tenfold decrease in [3H]-heparin binding capacity (Bmax = 1.9 x 10(6) sites per cell) compared to control cultures (Bmax = 1.7 x 10(7) sites per cell), which was irreversible after growing the cells for two additional passages in heparin-free medium. We also investigated protein kinase C (PKC) in HT-SMC in terms of both enzymatic activity and protein expression (evaluated by [3H]-staurosporine and [3H]-phorbol-12,13-dibutyrate binding). We found that HT-SMC had only half the PKC activity and expression as control SMC. Therefore, long-term treatment of rat aortic SMC with heparin allowed the selection of a less differentiated subpopulation of cells, exhibiting low sensitivity to the growth inhibition by heparin, which could be related to the low capacity of binding heparin and to a lower PKC activity and/or expression.
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PMID:Characterization of rat aortic smooth muscle cells resistant to the antiproliferative activity of heparin following long-term heparin treatment. 804 Jan 84

The present study was designed to examine whether heparin inhibits basal or stimulated endothelin-1 production by arginine vasopressin (AVP) and platelet-derived growth factor (PDGF) in cultured rat mesangial cells. In addition, the reversibility of the heparin effect on mesangial cell endothelin-1 production was examined. AVP and PDGF stimulated endothelin-1 secretion in a concentration-dependent manner in these cells. Heparin (10 to 100 U/ml) exhibited concentration-related inhibition of AVP- and PDGF-stimulated endothelin-1 secretion. Heparin also had weak but significant inhibitory effects on basal endothelin-1 secretion in these cells. The protein kinase (PKC)-activating phorbor ester, phorbor myristate acetate (PMA), stimulated endothelin-1 secretion and heparin inhibited PMA-stimulated endothelin-1 secretion. In addition, the inhibitory effect of heparin was completely abolished in PKC-depleted mesangial cells. Mesangial cells which were exposed to a high concentration (100 U/ml) of heparin for 24 hours were capable of producing endothelin-1 after a short lag period of removal of heparin from the culture medium. These mesangial cells also showed recovery of responses to AVP and PDGF by secreting a significantly greater amount of endothelin-1 than the non-stimulated level. These results indicate that heparin potently inhibits mesangial cell endothelin-1 production, especially when stimulated by AVP or PDGF. This inhibitory effect of heparin is probably PKC dependent, and reversible.
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PMID:Heparin inhibits endothelin-1 production in cultured rat mesangial cells. 812 2

Esophageal circular muscle cells isolated by enzymatic digestion contracted in response to acetylcholine (ACh) and in response to the protein kinase C (PKC) agonist 1,2-dioctanoylglycerol (1,2 DAG). Both responses were blocked by PKC antagonists but not by calmodulin antagonists. Furthermore, specific PKC activity, measured in the particulate fraction of the muscle, increased in response to cholinergic stimulation, suggesting that ACh-induced contraction is mediated by a PKC-dependent pathway. ACh-induced contraction decreased with decreasing extracellular Ca2+ and was blocked in Ca(2+)-free physiological salt solution (PSS). Similarly, contraction by the nonhydrolyzable GTP analogue guanosine 5'-O-(3-thiotriphosphate) was blocked by removal of Ca2+ from the PSS. Diacylglycerol production in response to ACh was reduced when extracellular Ca2+ was reduced from 2 to 0.5 mM and was abolished in Ca(2+)-free PSS. The response to 1,2-DAG, however, did not significantly change as extracellular Ca2+ or cytosolic Ca2+ was reduced to zero. Heparin (10 micrograms/ml), thapsigargin (3 microM), or the Ca2+ ionophore A-23187 (3 microM) had no effect on 1,2-DAG or ACh-induced contraction in permeable cells. The data suggest that contraction in response to ACh is mediated by influx of extracellular Ca2+ and a PKC-dependent pathway. Ca2+ may be required mainly to activate the phospholipases responsible for production of diacylglycerol, since contraction of esophageal muscle cells in response to 1,2-DAG is Ca2+ independent.
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PMID:Calcium requirements for acetylcholine-induced contraction of cat esophageal circular muscle cells. 814 7

Heparin inhibits the migration and proliferation of arterial smooth muscle cells and modifies the extracellular matrix. These effects may be the result of heparin's effects on proteinases that degrade the matrix. We have previously reported that heparin inhibits the induction of tissue-type plasminogen activator and interstitial collagenase mRNA. We have investigated the possibility that heparin affects other members of the matrix metalloproteinase family. Phorbol ester increased the levels of mRNA of collagenase, 92-kD gelatinase and stromelysin as well as the synthesis of these proteins. These effects were inhibited by heparin, but not by other glycosaminoglycans, in a dose-dependent manner. The induction of these matrix metalloproteinases was also inhibited by staurosporine and pretreatment with phorbol ester indicating the involvement of the protein kinase C pathway. In contrast, the 72-kD gelatinase was expressed constitutively and was not affected by phorbol ester or heparin. Tissue inhibitor of metalloproteinase-1 was expressed constitutively and was slightly increased by phorbol ester. It was not affected by heparin. Thus, heparin inhibits the production of four proteinases (tissue plasminogen activator, collagenase, stromelysin and 92-kD gelatinase) that form an interdependent system capable of degrading all the major components of the extracellular matrix.
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PMID:Heparin inhibits the induction of three matrix metalloproteinases (stromelysin, 92-kD gelatinase, and collagenase) in primate arterial smooth muscle cells. 818 30


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