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)

Balb/MK keratinocytes require epidermal growth factor for proliferation and terminally differentiate in response to elevated extracellular Ca2+ concentrations. The molecular pathways controlling cell differentiation in this system have yet to be established. We show that a dramatic and sustained activation of phosphoinositide metabolism is produced upon addition of Ca2+ to Balb/MK cultures. The pattern of inositol trisphosphate isomers released in response to Ca2+ challenge appeared to be atypical. Inositol 1,3,4-trisphosphate release was observed by 30s and was produced earlier than any alteration in inositol 1,4,5-trisphosphate levels. Concomitant with the liberation of inositol phosphates, an increased production of diacylglycerol was observed. Despite a 3-fold increase in diacylglycerol levels detected even at 12 h after Ca2+ addition, no evidence of functional activation or down-regulation of protein kinase C was found. This was established by measuring p80 phosphorylation, epidermal growth factor binding, and protein kinase C levels by immunoblotting. Analysis of the diacylglycerol generated following Ca2+ addition to Balb/MK cells revealed that a significant proportion of that lipid was an alkyl ether glyceride molecular species. Therefore, it is possible that this diacylglycerol molecular species may play a role in the Ca2+-induced differentiation program of Balb/MK cells through mechanisms other than stimulation of classical protein kinase C.
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PMID:The calcium signal for Balb/MK keratinocyte terminal differentiation induces sustained alterations in phosphoinositide metabolism without detectable protein kinase C activation. 247 92

An 80-kDa protein (p80), previously reported to be a major protein kinase C substrate in preneoplastic JB6 mouse epidermal cells, has been shown to be transiently phosphorylated by phorbol 12-O-tetradecanoate 13-acetate. Phosphorylation was maximal at 2 hr of phorbol 12-O-tetradecanoate 13-acetate treatment and returned to basal levels by 24 hr. In contrast, using a p80-specific antibody, we found that phorbol 12-O-tetradecanoate 13-acetate treatment produced no increase in p80 concentration. p80 showed a progressive decrease in JB6 cells during progression from a preneoplastic to neoplastic phenotype. The lack of p80 expression in neoplastic cells was not attributable to lack of protein kinase C; the protein kinase activity and protein concentration were similar in cells of all three phenotypes. When p80 mRNA was analyzed by hybridization to a putative p80 cDNA clone, its relative concentration paralleled that of p80 protein, with high levels present in preneoplastic JB6 cells, and little or no evidence for p80-hybridizing RNA in transformed cells. Thus, p80 appears to be regulated pretranslationally at the level of mRNA concentration during preneoplastic progression in mouse epidermal JB6 cells.
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PMID:Differential expression of an 80-kDa protein kinase C substrate in preneoplastic and neoplastic mouse JB6 cells. 279 14

Recently the cDNA for two different forms of TNF receptor, with gene products of molecular masses of 60 and 80 kDa, have been cloned. In the present report, we investigated the effects of phorbol ester and dibutyryl cAMP on the regulation of the transcript for each type of TNF receptor in U-937 cells. Our results indicate that exposure of these cells to either phorbol ester or dibutyryl cAMP increases the steady state mRNA levels of the 80 kDa form. This effect is dose- and time-dependent. The induction of the p80 receptor transcript by PMA and dibutyryl cAMP was additive suggesting independent mechanisms of induction. Under identical conditions, both agents failed to induce the transcript for the p60 form of the TNF receptor. As demonstrated by actinomycin D pulse-chase experiment, the mRNA for the p80 receptor was found to be highly stable with an approximate half-life of 16 h. No significant change in the half-life was observed when cells were treated with phorbol ester. The mechanisms by which phorbol ester and dibutyryl cAMP induce the upregulation of p80 receptor mRNA appear to be different. Induction of receptor transcript by cycloheximide suggests the presence of a labile repressor protein. Interestingly, the effect of cycloheximide on the induction of the p80 mRNA was found to be additive with that of dibutyryl cAMP but not with phorbol ester. 1-(5-Isoquinolinylsufonyl)-2-methylpiperazine (H7) and N[2-(methylamino) ethyl]-5-isoquinolinesulfonamide (H-8), inhibitors of protein kinase C and protein kinase A, respectively, both inhibited the phorbol ester-mediated induction of the p80-transcript but not that mediated through dibutyryl cAMP. Since dibutyryl cAMP undergoes intracellular dissociation into cAMP and butyric acid, we found that exposure of cells to sodium butyrate alone could induce p80 mRNA in a dose-dependent manner, thus suggesting the role of histone hyperacetylation. Furthermore forskolin treatment, an intracellular inducer of cAMP, increased the receptor transcript level whereas isobutylmethylxanthine, an inhibitor of phosphodiesterase, had no effect. Interestingly, while the p80 form of the TNF receptor mRNA levels was elevated by both phorbol ester and dibutyryl cAMP, only dibutyryl cAMP increased the TNF binding; phorbol ester treatment decreased the binding activity. Thus, our results demonstrate that the genes for the two forms of TNF receptors are differentially regulated. Furthermore, the mechanism of regulation by PMA differs from that by dibutyryl cAMP.
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PMID:Regulation of two forms of the TNF receptors by phorbol ester and dibutyryl cyclic adenosine 3',5'-monophosphate in human histiocytic lymphoma cell line U-937. 768 79

Incubation of B chronic lymphocytic leukemia (B-CLL) cells with phorbol esters resulted in the phosphorylation of three Triton-soluble, heat-stable, acidic proteins with apparent M(r) of 80 KDa, 60 KDa and 43 KDa. The characteristics of the three proteins suggested that they could be related to the myristoylated, alanine-rich, C-kinase substrate (MARCKS). p80 was immunoprecipitated with an antibody against the N-terminal peptide of MARCKS. p43 co-migrated with mouse MRP/Mac-MARCKS (MARCKS-related protein). p60 is the most prominent substrate of protein kinase C in B-CLL cells.
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PMID:Phosphorylation of the MARCKS family of protein kinase C substrates in human B chronic lymphocytic leukemia cells. 776 46

Staurosporine, an inhibitor of protein kinase C, is commonly used to inhibit the growth factor-induced signal transduction pathway at the post-receptor level. In this report, we examined the effect of staurosporine on the constitutive expression of tumor necrosis factor (TNF) receptors in K562, a human erythroblastoid leukemic cell line. Exposure of these cells to staurosporine enhanced cell surface expression of TNF receptors by almost 7-fold in a dose- and time-dependent fashion. Maximum induction occurred at a concentration of 20 nM of the agent for 16 h at 37 degrees C. Induction of the TNF receptor was found to be temperature-dependent. No induction was observed at 22 or at 4 degrees C, suggesting the role of cell metabolism. Scatchard analysis indicated an increase in receptor number without any change in receptor affinity. TNF receptors were induced by staurosporine on a wide variety of human cells of both epithelial (primarily p60 receptors) and myeloid (mainly p80 receptor) origin. Receptor-specific antibodies showed that both TNF receptors were induced. The induction was abolished by inhibitors of protein synthesis, thus suggesting the de novo synthesis of the receptor. Furthermore, we found that staurosporine had no effect on the internalization or shedding of the receptor, but it induced the mRNA for both forms of the TNF receptor. Inhibitors of tyrosine kinases had no effect on the induction of TNF receptors. Modulation of the receptor number by staurosporine correlated with the enhancement of antiproliferative effects of TNF against different tumor cells. Thus, overall these results indicate that protein kinase C may be involved in the signal transduction of TNF not only at the postreceptor level but also at the receptor level.
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PMID:Staurosporine induces the cell surface expression of both forms of human tumor necrosis factor receptors on myeloid and epithelial cells and modulates ligand-induced cellular response. 814 8

The phorbol ester, 12-deoxyphorbol-13-O-phenylacetate-20-acetate (DOPPA) has been shown to activate specifically the protein kinase C (PKC)-beta 1 isozyme in vitro (1). We have investigated the potential of DOPPA as a PKC-beta 1/2 isozyme-specific agonist in intact cells, employing U937 cells, which express beta 1/2, epsilon and zeta PKC and in Swiss 3T3 cells which lack PKC-beta 1/2 but express alpha, delta, epsilon and zeta PKC. Immunoblot analysis with isozyme-specific antibodies indicated that DOPPA can mediate the subcellular redistribution and down-modulation of all endogenous PKC isozymes (except PKC-zeta) in both U937 and Swiss 3T3 cells. Prolonged treatment (> 6 h) of cultures in down-modulation studies is complicated by the metabolism of DOPPA to 12-deoxyphorbol-13-phenylacetate (DOPP), a compound which activates all PKC isozymes tested in vitro (Ryves, W. J., et al. (1991) FEBS Lett., 288, 5-9). Nevertheless, because DOPPA induced rapid and dose-dependent phosphorylation of p80 in cells which do not express PCK-beta, p80 phosphorylation in Swiss 3T3 cells indicates that DOPPA can activate a non-beta PKC in vivo. The data suggest that DOPPA cannot be used as a PKC-beta-selective agonist in intact cell studies.
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PMID:12-Deoxyphorbol-13-O-phenylacetate-20-acetate is not protein kinase C-beta isozyme-selective in vivo. 831 25

Protein kinase C modulates the receptor for tumor necrosis factor (TNF) in wide variety of different cell types. However, there is no information about the role of phosphatases in the regulation of the TNF receptor. In this report, we investigated the effect of okadaic acid, an inhibitor of serine/threonine phosphatases, on TNF receptors in U-937 cells, a human histiocytic lymphoma cell line. In our study okadaic acid induced a dose- and time-dependent down-modulation of TNF receptors. On exposure of cells to 0.5 microM okadaic acid for 60 min at 37 degrees C, a complete down-regulation of the receptors was observed, but no modulation occurred at 4 degrees C. Scatchard analysis of the binding data on U-937 cells revealed that okadaic acid caused a decrease in the high affinity cell surface receptor number without a significant change in the affinity constant. The down-regulation of the TNF receptor by okadaic acid was not specific to U-937 cells, as it was also observed with several other cell types. Okadaic acid had no significant effect on the internalization of the receptor, but it did induce a shedding of the TNF receptor from its cell surface. On exposure of cells to okadaic acid, a dose- and time-dependent increase in the 40-kDa polypeptide was detected in the medium with anti-p80 antibodies by Western blot analysis. The secreted product was also found to bind TNF. The mechanism by which okadaic acid down-modulates the TNF receptor appears to be quite different from that of phorbol ester. First, okadaic acid synergistically potentiated the effect of phorbol ester. Second, the phorbol ester-mediated down-modulation could be blocked by H-7 and staurosporine, well known protein kinase C inhibitors, but these inhibitors had no effect on okadaic acid-mediated response. Third, phorbol ester-mediated down-modulation of the TNF receptor was reversible after removal of the agent, but the response mediated through okadaic acid was irreversible. Fourth, the TNF receptor was down-regulated by okadaic acid but not by phorbol ester in cells depleted of protein kinase C. Thus, overall, our results demonstrate that serine/threonine phosphatases can down-modulate and induce shedding of TNF receptors by a mechanism distinct from that of protein kinase C.
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PMID:Okadaic acid induces down-modulation and shedding of tumor necrosis factor receptors. Comparison with another tumor promoter, phorbol ester. 838 75

We have shown previously that novel protein kinase Cepsilon (nPKCepsilon) plays a key role in the basal and thyrotropin-releasing hormone (TRH)-stimulated prolactin (PRL) secretion in rat pituitary GH4C1 cells (Akita et al., J. Biol. Chem. 1994, 269, 4653-4660). Here we examined the region downstream of nPKCepsilon activation in order to understand the molecular mechanism by which nPKCepsilon mediates TRH-induced signal transduction. Exposure of GH4C1 cells to TRH causes a stimulation of the phosphorylation of a p80 (Mr approximately 80 000, pI approximately 4.3) and two p19 (p19a and b; Mr approximately 19 000, pI approximately 5.6 and 5.5, respectively). Phorbol ester, a potent activator of protein kinase C (PKC), also enhances these phosphorylations, whereas bisindolylmaleimide I, a specific inhibitor of PKC, clearly inhibits the phosphorylation of p80. p80 and p19 were identified as myristoylated alanine-rich C kinase substrate (MARCKS) and stathmin, respectively, as assessed by their two-dimensional gel electrophoretic profiles and their stabilities to heat and acid treatment. In nPKCepsilon-overexpressing stable clones, the phosphorylated level of MARCKS but not stathmin was high in the resting state, and enhanced and sustained upon TRH stimulation, correlating with the increased activation of nPKCepsilon. TRH stimulates the release of MARCKS from the membrane/cytoskeletal fraction to the cytosol fraction. These results, taken together with previous data concerning PRL secretion, suggest that MARCKS, a regulatory component of the cytoskeletal architecture, is the major substrate of nPKCepsilon in vivo, and that its phosphorylation may regulate TRH-stimulated PRL secretion.
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PMID:Involvement of protein kinase C epsilon in thyrotropin-releasing hormone-stimulated phosphorylation of the myristoylated alanine-rich C kinase substrate in rat pituitary clonal cells. 1067 27

Clinical and experimental studies have implicated high circulating levels of the cytokine tumour necrosis factor-alpha (TNF-alpha) in the pathogenesis of insulin resistance, not only in obesity and diabetes, but also in clinical conditions associated with cachexia and sepsis. TNF-alpha impairs insulin-mediated glucose uptake in adipocytes, but because of lipolytic effects the interpretation of clinical studies and the extent to which TNF-alpha affects muscle insulin sensitivity are unclear. In addition, protein kinase C (PKC) has recently been implicated in the mechanism of TNF-alpha-induced insulin resistance. The present study investigated the effects of TNF-alpha and a PKC inhibitor (RO-318220) on basal and insulin-stimulated 2-[(3)H]deoxyglucose uptake in cultured L6 myoblasts. Reverse transcriptase-PCR analysis confirmed that L6 myoblasts express TNF-alpha receptors I and II (p60 and p80). Dose-response curves for glucose uptake were fitted to a quadratic function to derive C(I-150) values (concentration of insulin required to increase glucose uptake by 50%). Incubation with TNF-alpha at 1 or 10 ng/ml for 24 h had no significant effect on basal glucose uptake, insulin sensitivity or maximal insulin responsiveness. C(I-150) values (means+/-S.E.M.) were as follows: basal, 91.2+/-13 nM; 1 ng/ml TNF-alpha, 102+/-12 nM; and basal, 70.8+/-13 nM; 10 ng/ml TNF-alpha, 43.7+/-40 nM. PKC inhibition markedly attenuated glucose uptake, but there was no difference in insulin sensitivity with RO-318220 alone compared with RO-318220+TNF-alpha. In conclusion, although increased TNF-alpha expression and plasma concentrations have been implicated in the pathogenesis of insulin resistance in various clinical states, there is no evidence that TNF-alpha impairs insulin-stimulated glucose uptake in a skeletal-muscle-derived cell line.
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PMID:Effects of tumour necrosis factor-alpha and inhibition of protein kinase C on glucose uptake in L6 myoblasts. 1099 95

The plasma membrane is not homogeneous but contains specific subcompartments characterized by their unique lipid and protein composition. Based on their enrichment in various signaling molecules, these membrane microdomains are recognized to be sites of localized signal transduction for a number of extracellular stimuli. We have previously shown that fibroblast growth factor-2 (FGF2) induced a specific signaling response within a lipid raft membrane microdomain in human neuroblastoma cells characterized by the tyrosine phosphorylation of a p80 phosphoprotein. Herein, we show that this protein is the signaling adaptor FRS2 and that it is localized exclusively to lipid rafts in vitro and in vivo. We have examined how the tyrosine phosphorylation and serine-threonine phosphorylation of FRS2 within lipid rafts affect the response of cells to FGF2 signaling. Our data suggest that activation of protein kinase C, Src family kinases, and MEK1/2 are involved in regulating serine-threonine phosphorylation of FRS2, which can indirectly affect FRS2 phosphotyrosine levels. We also show that Grb2 is recruited to lipid rafts during signaling events and that activation of MEK1/2 by different mechanisms within lipid rafts may lead to different cellular responses. This work suggests that compartmentalized signaling within lipid rafts may provide a level of specificity for growth factor signaling.
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PMID:Fibroblast growth factor-2-induced signaling through lipid raft-associated fibroblast growth factor receptor substrate 2 (FRS2). 1257 Dec 52


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