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

Most platelet agonists activate and elevate the cytosolic free calcium concentration in human platelets through receptor-dependent mechanisms that are antagonized by cAMP- and cGMP-elevating agents. Nitrovasodilators such as nitroprusside and endothelium-derived relaxing factor are potent cGMP-elevating platelet inhibitors. In the present study, the role of cGMP and cGMP-dependent protein kinase in nitrovasodilator inhibition of ADP- and thrombin-evoked calcium elevation and activation of human platelets was investigated. Preincubation of platelets with 8-(4-chlorophenylthio)guanosine 3',5'-cyclic monophosphate (8-pCPT-cGMP; a membrane-permeant selective activator of the cGMP-dependent protein kinase that does not significantly affect cGMP-regulated phosphodiesterases) inhibited the thrombin-induced phosphorylation mediated by myosin light chain kinase and protein kinase C. Nitrovasodilator-induced protein phosphorylation in human platelets was distinct from that induced by cAMP-elevating prostaglandins and could be mimicked by 8-pCPT-cGMP. Preincubation of human platelets with nitrovasodilators or 8-pCPT-cGMP inhibited the ADP- and thrombin-evoked calcium elevation in the presence and absence of external calcium. Nitrovasodilators and 8-pCPT-cGMP also inhibited the agonist-induced Mn2+ influx, but stopped-flow experiments indicated that the ADP receptor-operated cation channel was not significantly inhibited. These results suggest that in human platelets nitrovasodilators inhibit the agonist-induced calcium mobilization from intracellular stores and the secondary store-related calcium influx but not the ADP receptor-operated cation channel. The results also suggest that these nitrovasodilator effects are mediated by cGMP and the cGMP-dependent protein kinase.
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PMID:Role of cGMP and cGMP-dependent protein kinase in nitrovasodilator inhibition of agonist-evoked calcium elevation in human platelets. 131 May 37

Diacylglycerol kinase activity was demonstrated in highly purified plasma membranes isolated from shoots and roots of dark-grown wheat (Triticum aestivum L.) by aqueous polymer two-phase partitioning. The active site of the diacylglycerol kinase was localized to the inner cytoplasmic surface of the plasma membrane using isolated inside-out and right-side-out plasma membrane vesicles from roots. The enzyme activity in plasma membrane vesicles from shoots showed a broad pH optimum around pH 7. The reaction was Mg2+ and ATP dependent, and maximal activity was observed around 0.5 mM ATP and 3 mM MgCl2. The Mg2+ requirement could be substituted only partially by Mn2+ and not at all by Ca2+. The phosphorylation of endogenous diacylglycerol was strongly inhibited by detergents indicating an extreme dependence of the lipid environment. Inositol phospholipids stimulated the activity of diacylglycerol kinase in plasma membranes from shoots and roots, whereas the activity was inhibited by R59022, a putative inhibitor of several diacylglycerol kinase isoenzymes involved in uncoupling diacylglycerol activation of mammalian protein kinase C.
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PMID:Diacylglycerol kinase in plasma membranes from wheat. 131 Aug 76

Effects of protein kinase C (PKC) on bradykinin (BK)-induced intracellular calcium mobilization, consisting of rapid Ca2+ release from internal stores and a subsequent sustained Ca2+ inflow, were examined in Fura-2-loaded osteoblast-like MC3T3-E1 cells. The sustained Ca2+ inflow as inferred with Mn2+ quench method was blocked by Ni2+ and a receptor-operated Ca2+ channel blocker SK&F 96365, but not by nifedipine. The short-term pretreatment with phorbol 12-myristate 13-acetate (PMA), inhibited BK-stimulated Ca2+ inflow, and the prior treatment with PKC inhibitors, H-7 or staurosporine, enhanced the initial internal release and reversed the PMA effect. Moreover, 6 h pretreatment with PMA caused similar effect on the BK-induced inflow to that obtained with PKC inhibitors, whereas 24 h pretreatment was necessary to affect the internal release. On the other hand, the translocation and down-regulation of PKC isozymes were examined after PMA treatment of MC3T3-E1 cells by immunoblot analyses of PKCs with the isozyme-specific antibodies. 6 h treatment with PMA induced down-regulation of PKC beta, whereas longer treatment was needed for down-regulation of PKC alpha. Taken together, it was suggested that the BK-induced initial Ca2+ peak and the sustained Ca2+ inflow through the activation of a receptor-operated Ca2+ channel, are differentially regulated by PKC isozymes alpha and beta, respectively, in osteoblast-like MC3T3-E1 cells.
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PMID:Different protein kinase C isozymes could modulate bradykinin-induced extracellular calcium-dependent and -independent increases in osteoblast-like MC3T3-E1 cells. 132 Apr 58

Microtubule-associated protein 2 (MAP2) kinase has been isolated and characterized from rat brain. The enzyme has an apparent M(r) of approximately 42,000 and its pI is 4.9. MAP2 was the preferred substrate, but it also phosphorylated myelin basic protein (MBP), histone V-S, tubulin and the PC12 protein substrate pp250. The enzyme is distinct from protein kinase C, cAMP-dependent kinase and the calcium/calmodulin-dependent kinases, as specific inhibitors of these kinases did not affect MAP2 phosphorylation. The addition of the relatively non-specific protein kinase inhibitor H7 (20 microM) had a modest inhibitory effect. The enzyme was active in both 5 mM Mn2+ and Mg2+, and displayed Kms for MAP2, MBP, and ATP of 56 nM, 254 nM, and 4 microM, respectively. This enzyme, which represents a low abundance protein in whole brain, is analogous to the MAP2 kinase observed in growth factor-stimulated cell lines.
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PMID:Isolation and characterization of microtubule-associated protein 2 (MAP2) kinase from rat brain. 132 16

A cytosolic insulin-sensitive serine kinase has been purified to apparent homogeneity in parallel from livers of control or acutely insulin-treated rats. The kinase is labile and requires rapid purification for stability. The kinase migrates as a band of apparent Mr = 90,000 on denaturing gels and elutes as a monomer on Superose 12 gel filtration. After sodium dodecyl sulfate-polyacrylamide gel electrophoresis and renaturation, the 90-kDa band presumed to be the kinase shows kinase activity toward myelin basic protein in situ. Substrates of the kinase include Leu-Arg-Arg-Ala-Ser-Leu-Gly (Kemptide), ribosomal protein S6, S6 peptide, a proline-rich peptide substrate, microtubule-associated protein 2, and myelin basic protein. The kinase also phosphorylates histones H1 and H2B, but does not autophosphorylate to a significant stoichiometry. The activity of the kinase is inhibited by fluoride, glycerophosphate, p-nitrophenyl phosphate, p-nitrophenol, heparin, quercetin, poly-L-lysine, and potassium phosphate, but is unaffected by calcium, cAMP, spermine, protein kinase inhibitor peptide, phorbol myristate acetate, calcium plus phosphatidylserine, or vanadate. The kinase will utilize magnesium (10 mM) as well as manganese (1 mM) as a cofactor for maximal phosphotransferase activity. The kinase is not detected by immunoblotting with antibodies directed against protein kinase C or type II S6 kinase. Taken together, these properties distinguish this kinase from other insulin-sensitive kinases that have been described previously. The purified kinase from livers of insulin-treated rats shows a 5-20-fold higher specific activity compared to enzyme prepared from control rats, suggesting a covalent modification as the mechanism of activation. Incubation of purified, insulin-stimulated kinase with purified phosphatase 2A leads to deactivation of the kinase activity, and the phosphatase inhibitor nitrophenyl phosphate blocks this deactivation. The insulin-activated kinase fails to immunoblot with anti-tyrosine phosphate antibodies. Taken together, these results indicate that insulin activates this novel cytosolic protein kinase by a mechanism that causes its phosphorylation on serine or threonine residues.
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PMID:Purification and characterization of a cytosolic insulin-stimulated serine kinase from rat liver. 153 38

Human platelets were loaded with the fluorescent Na(+)-sensitive dye sodium-binding benzofuran isophtalate (SBFI), and changes in the fluorescence excited at 345 and 385 nm were analyzed after manipulations that evoked predictable changes in the cytosolic Na+ concentration ([Na+]i). Raising [Na+]i by either gramicidin D or monensin specifically increased the fluorescence excited at 345 nm and decreased that excited at 385 nm. Hence, calculation of changes in the 345/385 nm excitation ratio yields an estimate of actual changes in [Na+]i. A transient activation of Na+/H+ exchange evoked by addition of acidified platelets to buffer, pH 7.4, evoked a transient rise in [Na+]i. The re-establishment of basal [Na+]i could be prevented by ouabain, indicating an involvement of the Na+,K(+)-ATPase. Upon stimulation by 0.5 unit/ml of thrombin, [Na+]i immediately increased by 16 +/- 4 mM and this rise continued for at least 60 min after addition of agonist, albeit at a lower rate. This latter sustained rise could not be curtailed by scavenging thrombin by means of hirudin. Addition of ouabain or the phorbol ester 12-O-tetradecanoylphorbol-13-acetate induced a comparable slow rise in the 345/385 excitation ratio. This may indicate a protein kinase C-mediated inhibition by thrombin of the Na+,K(+)-ATPase. In the absence of extracellular Ca2+ (Ca2+o), the [Na+]i gain was augmented to 38 +/- 9 mM. This additional uptake of Na+ was prevented by (i) Mn2+ ions, (ii) La3+ ions, (iii) the blocker of receptor-mediated Ca2+ entry (1-[beta[3-(4-methoxyphenyl)propoxyl]-4-methoxyphenethyl]-1H-im ida zole hydrochloride), and (iv) by hirudin which reversed receptor occupancy by thrombin. These findings suggest that the additional thrombin-induced [Na+]i gain in the absence of Ca2+o is due to Na+ influx through a Ca2+ entry pathway. The increase in [Na+]i in the presence of Ca2+o results from Na+ influx via Na+/H+ exchange.
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PMID:Further characterization of the mechanisms mediating the rise in cytosolic free Na+ in thrombin-stimulated platelets. Evidence for inhibition of the Na+,K(+)-ATPase and for Na+ entry via a Ca2+ influx pathway. 164 80

Engagement of membrane IgM on a number of human and murine B-cell lines induced activation of a Mn(2+)-preferring serine/threonine kinase that phosphorylated microtubule-associated protein-2 (MAP-2) in vitro. B-cell MAP-2 kinase (MAP-2K) activity could be fractionated into two peaks by sequential DEAE and hydrophobic chromatography. Although peak I included two tyrosine phosphoproteins of molecular mass 36 and 38 kDa, peak II showed a single 42-kDa tyrosine phosphoprotein (pp42). Since all kinase activity could be removed from peak II material over an antiphosphotyrosine immune affinity column, it suggests that pp42 is identical with lymphoid MAP-2K. Although peak I activity showed a similarity to peak II with regard to its preference for Mn2+, sensitivity to phosphatase exposure, and resistance to a range of common serine kinase inhibitors, it is not clear whether these activities are related. MAP-2 kinase activity could also be induced by treatment with the phorbol ester, phorbol myristate 13-acetate, suggesting that protein kinase C may also be involved with MAP-2K regulation. Although MAP-2K activity reached a peak response within minutes of receptor ligation, there were differences in the rates of dephosphorylation of pp42 and decline of MAP-2K activity in different B-cell lines. The tyrosine phosphatase inhibitor, vanadate, transformed a rapidly reversible MAP-2K response in BAL 17.2 cells into a sustained state of activation that resembled the kinetics of activation in WEHI-231 cells. The latter finding implies involvement of a tyrosine phosphatase, which opposes the effect of an inducing tyrosine kinase.
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PMID:Stimulation of B-cells via the membrane immunoglobulin receptor or with phorbol myristate 13-acetate induces tyrosine phosphorylation and activation of a 42-kDa microtubule-associated protein-2 kinase. 165 69

Isopeptides of the endothelin gene family, including sarafotoxin, caused a concentration-dependent increase in intracellular-free calcium (cytosolic calcium) in cultured human and rat glomerular mesangial cells. The calcium increments had two components, a spike increment resulting from mobilization of intracellular calcium and a sustained calcium increment as a result of endothelin activation of calcium influx. ET-dependent calcium influx occurred through dihydropyridine-insensitive calcium channels which were blocked by NiCl and allowed entry of Mn2+. In human mesangial cells, ET stimulated periodic oscillations of cytosolic Ca2+ that reflect synchronous signalling in localized populations. Calcium signalling evoked by the ET isopeptides showed cross-desensitization among the four ET-peptides tested. Although protein kinase C activation acutely reduced ET-induced calcium signalling, the desensitization by ET isopeptides was independent of protein kinase C. If ET was added to cells incubated with an inhibitor of the endoplasmic reticulum calcium ATPase, it became apparent that ET activated a calcium efflux pathway which reduced cytosolic calcium. These data, therefore, demonstrate that calcium signalling is a common response in both human and rat cultured mesangial cells incubated with different ET isopeptides.
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PMID:The effects of endothelin on cytosolic calcium in cultured human and rat glomerular mesangial cells. 166 3

Endothelin (ET)-related peptides robustly stimulated [3H]-inositol phosphate (IP) formation in cultured cerebellar granule cells, astrocytes, and C6 glioma cells. Their agonist selectivities were ET-1 = ET-2 greater than or equal to sarafotoxin S6b greater than ET-3 greater than big ET-1 for granule cells and ET-1 greater than or equal to ET-2 greater than or equal to S6b greater than big ET-1 greater than ET-3 for cerebellar astrocytes and C6 glioma cells. These effects were Ca(2+)-dependent but insensitive to antagonists of L-type Ca2+ channels and the Na+/Ca2+ antiporter. Pretreatment of cells with ET-1 or S6b induced homologous desensitization of phosphoinositide (PI) response mediated by ET receptors. Long-term pertussis toxin (PTX) treatment attenuated the phosphoinositide (PI) response in astrocytes and glioma but not in granule cells. ET-1 and its related peptides increased [Ca2+]i in C6 glioma by two distinct pathways: IP3-induced Ca2+ mobilization or receptor-operated Ca2+ influx. La3+, Mn2+, and Cd2+ inhibited the Ca2+ influx and sustained PI turnover, while Ca2+ mobilization was attenuated by phorbol ester and TMB-8. ET-induced Ca2+ influx was essential for the sustained [Ca2+]i increase and PI turnover. Homologous desensitization of [Ca2+]i increase was also noted. In cerebellar granule cells, ET evoked the release of [3H]D-aspartate from these neurons. This action appears to be dependent on PI hydrolysis and [Ca2+]i increase and modulated by protein kinase C.
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PMID:Endothelin-induced activation of phosphoinositide turnover, calcium mobilization, and transmitter release in cultured neurons and neurally related cell types. 172 40

The effects of phorbol ester (TPA) and other known stimulators such as tumor necrosis factor (TNF), interleukin-1, and lipopolysaccharide on induction of mRNA for manganese-superoxide dismutase (Mn-SOD) were investigated in various cell lines. TPA enhanced Mn-SOD mRNA expression in TNF-resistant cell lines including HeLa cells, in which the other reagents also induced expression of the gene, but did not affect TNF-sensitive cells, in which the other stimulators did not alter expression of the gene. HeLa cells which had been desensitized to TPA by pretreatment with TPA for 24 h expressed Mn-SOD mRNA at a slightly higher level than the cells without TPA treatment. TPA-pretreated cells stimulated with TNF, however, expressed Mn-SOD mRNA at about twice the level of TNF-stimulated, TPA-untreated cells. When protein synthesis was inhibited by cycloheximide during TPA pretreatment, TNF no more enhanced the Mn-SOD mRNA accumulation. These data suggest that at least two separate signal-transducing pathways are involved in expression of this gene. One is triggered by protein kinase C activation itself in the absence of new protein synthesis. The other can be activated by stimulation with TNF, interleukin-1, or lipopolysaccharide and in which a protein factor that can be induced by TPA treatment is involved.
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PMID:Phorbol ester induces manganese-superoxide dismutase in tumor necrosis factor-resistant cells. 174 13


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