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 expressions of the protooncogenes c-jun and jun D have been investigated in dog thyrocytes in a primary culture whose proliferation is stimulated by three distinct intracellular signaling pathways (1) the thyrotropin (TSH) or forskolin-cyclic-AMP-mediated cascade; (2) the protein kinase C pathway activated by diacylglycerol (DAG) and phorbol esters (TPA); (3) a protein tyrosine kinase system activated by epidermal growth factor (EGF). While the first cascade is compatible with the differentiated state of the cell, the two latter pathways induce dedifferentiation. Following the stimulation by TPA or EGF, the expression of c-jun was increased and the expression of jun D was faintly increased. Both expressions are superinduced in the presence of cycloheximide as in mitogenically stimulated fibroblasts but, in the presence of cycloheximide alone, the expressions of c-jun and jun D are clearly unstable with time. This indicates that cycloheximide controls should be included at all time points examined in such experiments. Increasing intracellular concentrations of cyclic-AMP by forskolin or TSH was followed by an inhibition of the expression of c-jun. This inhibition was independent of protein synthesis. Similarly, the TPA or EGF stimulation of c-jun expression was also inhibited by TSH or forskolin, as in fibroblasts in which cyclic-AMP inhibits proliferation. Our results show that the expression of c-jun is not universally correlated with the stimulation of cell proliferation. The stimulation of c-jun expression is not common between the three mitogenic pathways. It thus represents another of the very different responses elicited by the cyclic-AMP cascade as compared to the more studied tyrosine kinase and protein kinase C mitogenic pathways.
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PMID:Differential regulation of protooncogenes c-jun and jun D expressions by protein tyrosine kinase, protein kinase C, and cyclic-AMP mitogenic pathways in dog primary thyrocytes: TSH and cyclic-AMP induce proliferation but downregulate C-jun expression. 165 70

In a previous study, we found that addition of serum to confluent Clone 9 cells, a nontransformed rat liver cell line, increased the abundance of mRNA alpha 1 and mRNA beta 1 at 3 h by 2- and 2.7-fold, respectively [Bhutada et al. Am. J. Physiol. 258 (Cell Physiol. 27): C1044-C1050, 1990]. We now report that exposure of these cells to 160 nM 12-O-tetradecanoylphorbol 13-acetate (TPA) for 6 h increases mRNA alpha 1 and mRNA beta 1 by 1.7 +/- 0.2- and 2.1 +/- 0.3-fold, respectively. Incubation in the presence of 160 nM TPA for 24 h reduced high-affinity phorbol dibutyrate-binding sites [dissociation constant (Kd) = 5 nM; maximum binding (Bmax) = 1.2 pmol/mg protein] to undetectable levels. In such cells, exposure to 10% serum for 6 h still resulted in two- and fourfold increment in mRNA alpha 1 and mRNA beta 1 abundances, respectively, while further addition of TPA to these protein kinase C (PKC)-depleted cells resulted in no change in the subunit mRNA abundances. The increments in mRNA alpha 1 content in response to 10% serum and 160 nM TPA at 6 h were additive, whereas the increments in mRNA beta 1 were not. The following agents increased mRNA alpha 1 and mRNA beta 1 abundance in both control and PKC-depleted cells: epidermal growth factor, platelet-derived growth factor, basic fibroblast growth factor, insulin, dexamethasone, and hypothyroid calf serum. In contrast, N6,2'-O-dibutyryl-adenosine 3',5'-cyclic monophosphate and aldosterone had no effect.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Serum and growth factor induction of Na(+)-K(+)-ATPase subunit mRNAs in Clone 9 cells: role of protein kinase C. 165 70

Okadaic acid, a potent tumor promoter and inhibitor of phosphoserine/threonine protein phosphatases 1 and 2A, produces a large increase in epidermal growth factor (EGF) receptor phosphorylation in several cell types. The increases are limited to phosphoserine and phosphothreonine residues. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a distinct tumor promoter and protein kinase C activator, also induces serine/threonine phosphorylation of the EGF receptor and is known to modulate receptor functions. Comparison of okadaic acid and TPA influences on the EGF receptor show significant differences. Okadaic acid did not promote phosphorylation of Thr-654, a major site of TPA-induced phosphorylation. However, other sites of phosphorylation were similar for the two tumor promoters. In vitro experiments with purified protein phosphatase 2A demonstrate the insensitivity of Thr-654 phosphorylation, which regulates EGF receptor function, to dephosphorylation by this okadaic acid-sensitive protein phosphatase. In contrast to TPA, okadaic acid did not attenuate the tyrosine kinase activity or ligand binding capacity of the EGF receptor. However, okadaic acid did produce a decrease in EGF-stimulated inositol phosphate formation in a manner distinct from that of TPA.
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PMID:Okadaic acid-induced hyperphosphorylation of the epidermal growth factor receptor. Comparison with receptor phosphorylation and functions affected by another tumor promoter, 12-O-tetradecanoylphorbol-13-acetate. 165 56

In this study we describe the activation of a protein kinase which phosphorylates a peptide, T669, comprising amino acids 663-681 of the epidermal growth factor receptor and containing the phosphate acceptor site Pro-Leu-Thr669-Pro. In the human epidermoid carcinoma cell line KB, T669 kinase activity in cytosolic extracts peaked (up to 15-fold compared with basal levels) 15-30 min after addition of interleukin-1 (IL-1) and closely paralleled receptor occupancy with a half-maximally effective concentration of approximately 100 pM IL-1 alpha. IL-1 treatment elevated T669 kinase activity to a variable extent in selected fibroblast lines, the hepatoma cell line HepG2, and the murine thymoma EL4 6.1. An IL-1 receptor-negative EL4 variant and the B cell lines 70Z/3, CB23, and RPMI 1788 did not respond in this way. All of the cell lines except 70Z/3 showed increased levels of T669 kinase when treated with the protein kinase C activator phorbol myristate acetate and/or with epidermal growth factor. This finding is in agreement with a previous study (Countaway, J. L., Northwood, I. C., and Davis, R. J. (1989) J. Biol. Chem. 264, 10828-10835). Activators of protein kinase A did not mimic the ability of IL-1 to stimulate T669 kinase activity, nor did the protein kinase C inhibitor staurosporine abrogate the effect of IL-1. T669 kinase activity from IL-1-stimulated KB cells was partially purified by ion exchange, hydrophobic interaction, and size exclusion chromatography. The partially purified enzyme phosphorylated myelin basic protein, a characteristic substrate of microtubule-associated protein-2 kinase (MAP-2 kinase) and the peptide Arg-Arg-Arg-(Tyr-Ser-Pro-Thr-Ser-Pro-Ser)4 from RNA polymerase II. Western blotting of chromatographic fractions revealed that T669 kinase activity corresponded with two proteins of 43 and 45 kilodaltons which cross-reacted with antibodies raised against peptide sequences of rat extracellular signal-regulated kinase-1/microtubule-associated protein-2 kinase. T669 kinase activity was critically dependent on the presence of phosphatase inhibitors. Since both the 43- and 45-kDa proteins, immunoprecipitated from [32P]phosphate-labeled cells, demonstrated a dramatic increase in their levels of serine, threonine, and tyrosine phosphorylation after brief treatment with IL-1, we conclude that IL-1 modulates the activity of these extracellular signal-regulated kinase/microtubule-associated protein-2 kinases by altering the level of their phosphorylation.
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PMID:Interleukin-1 represents a new modality for the activation of extracellular signal-regulated kinases/microtubule-associated protein-2 kinases. 165 5

Fibroblast growth factor 5 (FGF-5) is a member of the fibroblast growth factor family with transforming potential. It has been found to be expressed in several human tumor cell lines, but nothing is known about expression of this growth factor in normal cells and its biological functions. Here we show that the FGF-5 gene is expressed in exponentially growing normal human fibroblasts. In quiescent fibroblasts, expression of FGF-5 is strongly induced by serum and several growth factors such as platelet-derived growth factor (PDGF), epidermal growth factor (EGF) and transforming growth factor alpha (TGF-alpha). This induction can be mediated by at least two different pathways involving protein kinase C or cAMP-dependent kinases. Since the effect is independent of de novo protein synthesis, FGF-5 represents the product of a primary response gene. In addition our data suggest that FGF-5 is mitogenic for human fibroblasts, indicating the existence of an FGF-5-mediated positive feedback in these cells which could amplify and prolong the cellular response to the initial stimulus.
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PMID:Fibroblast growth factor 5 proto-oncogene is expressed in normal human fibroblasts and induced by serum growth factors. 165 9

Previous studies from our laboratory have demonstrated that epidermal growth factor (EGF), induces intracellular alkalinization in chicken granulosa cells by activating a sodium-dependent and amiloride-sensitive Na+/H+ antiporter. In the present investigation we have examined the possible involvement of protein kinase C (PKC) in the regulation of intracellular pH (pHi) by EGF in chicken granulosa cells. Intracellular pH in granulosa cells obtained from the two largest preovulatory follicles was determined spectrofluorometrically using the dye 2',7'-bis-(carboxyethyl)-5(6)-carboxyfluorescein. The resting pHi was 6.81 +/- 0.01 (n = 30) when the extracellular pH and sodium concentration were 7.3 and 144 mM, respectively. 12-O-Tetradecanoyl-phorbol-13-acetate (TPA; 50-400 ng/ml) and 1-oleoyl-2-acetylglycerol (OAG; 1-75 micrograms/ml) mimicked the actions of EGF by inducing a concentration-dependent increase in pHi which reached a maximum of 0.25-0.30 pH units. 4 alpha-Phorbol 12,13-didecanoate, a phorbol ester with no tumor promoting activity had no effect on pHi. Cytosolic alkalinization was observed within 10 min of the addition of each agent and increased over the 60-min observation period. Like EGF-induced cytosolic alkalinization, the increases in pHi in response to TPA or OAG were dependent on the presence of sodium concentration and were inhibited by amiloride, an inhibitor of the Na+/H+ antiporter. The effects of EGF, TPA, and OAG were attenuated by the PKC inhibitors 5-isoquinolinylsulfonyl-2-methyl piperazine and trifluoperazine. Down-regulation of granulosa cell PKC by pretreatment with TPA (200 ng/ml) for 2.5 h inhibited EGF-, TPA-, and OAG-induced cytosolic alkalinization. The effects of maximally stimulatory concentrations of EGF and TPA on cytosolic alkalinization were not additive. The increases in pHi induced by TPA and OAG, but not by EGF, were dependent on the presence of extracellular Ca++. These studies suggest that the EGF-induced intracellular alkalinization in chicken granulosa cells involves a PKC-mediated activation of the Na+/H+ antiporter.
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PMID:Epidermal growth factor elevates intracellular pH in chicken granulosa cells by activating protein kinase C. 165 20

Treatment of normal human fibroblasts with epidermal growth factor (EGF) results in the rapid (0.5 min) and simultaneous tyrosine phosphorylation of the EGF receptor (EGFr) and several other proteins. An exception to this tyrosine phosphorylation wave was a protein (42 kDa) that became phosphorylated on tyrosine only after a short lag time (5 min). We identified this p42 kDa substrate as the microtubule-associated protein (MAP) kinase using a monoclonal antibody to a peptide corresponding to the C-terminus of the predicted protein (Science 249, 64-67, 1990). EGF treatment of human fibroblasts at 37 degrees C for 5 min resulted in the tyrosine phosphorylation of 60-70% of MAP kinase as determined by the percent that was immunoprecipitated with antiphosphotyrosine antibodies. Like other tyrosine kinase growth factor receptors, the EGFr is activated and phosphorylated at 4 degrees C but is not internalized. Whereas most other substrates were readily tyrosine phosphorylated at 4 degrees C, MAP kinase was not. When cells were first stimulated with EGF at 4 degrees C and then warmed to 37 degrees C without EGF, tyrosine phosphorylation of MAP kinase was again observed. Treatment of cells with the protein kinase C activator phorbol myristate acetate (PMA) also resulted in the tyrosine phosphorylation of MAP kinase, and again only at 37 degrees C. Tryptic phosphopeptide maps demonstrated that EGF and PMA both induced the phosphorylation of the same peptide on tyrosine and threonine. This temperature and PMA sensitivity distinguishes MAP kinase from most other tyrosine kinase substrates in activated human fibroblasts.
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PMID:Temperature-dependent tyrosine phosphorylation of microtubule-associated protein kinase in epidermal growth factor-stimulated human fibroblasts. 166 89

Staurosporine is a potent but nonselective inhibitor of protein kinase C (PKC) and blocks responses to 12-O-tetradecanoylphorbol-13-acetate (TPA) in several cell types in vitro. In cultured primary mouse keratinocytes, however, staurosporine fails to inhibit TPA-mediated keratinocyte maturation and itself elicits responses that are similar to TPA (T. Sako et al., Cancer Res., 48: 4646-4650, 1988). After exposure to 10 nM staurosporine for 24 h, essentially all keratinocytes undergo morphological differentiation, whereas 160 nM TPA induces this response in about 50% of epidermal cells. These concentrations of staurosporine and TPA cause a 4-5-fold induction of epidermal transglutaminase activity and cornified envelopes, both markers of the terminal stage of keratinocyte differentiation. Staurosporine, but not TPA, also induces morphological and biochemical maturation in 2 neoplastic mouse keratinocyte cell lines, 308 and SP-1. The ability of staurosporine to elicit the same responses as TPA suggested that it may be functioning paradoxically as a PKC agonist in intact keratinocytes. In support of this hypothesis, staurosporine induces ornithine decarboxylase activity, inhibits 125I-labeled epidermal growth factor binding, and induces expression of c-fos mRNA. Down-regulation of PKC by pretreatment of primary keratinocytes with 60 nM bryostatin partially blocks staurosporine-mediated induction of cornified envelopes and inhibition of 125I-labeled epidermal growth factor binding, implicating PKC in these responses. The ability of staurosporine to mimic and/or enhance certain responses to TPA suggests that this agent is acting as a functional PKC agonist in cultured keratinocytes.
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PMID:Staurosporine induces protein kinase C agonist effects and maturation of normal and neoplastic mouse keratinocytes in vitro. 167 84

Interleukin 1 or tumor necrosis factor alpha can cause a transient down-modulation of epidermal growth factor (EGF) binding to quiescent fibroblast monolayers; the effect results from a reduction in EGF receptor (EGF-R) affinity and appears to be mediated by a protein kinase C (PKC)-independent mechanism. Here we show transient increases in EGF-R serine/threonine phosphorylation which are temporally coordinated with the effects on EGF binding; we also demonstrate that the cytokine-mediated phosphorylations, unlike those caused by PKC activators, have little discernible effect upon intrinsic EGF-R-associated tyrosine kinase activity. Cytokine-mediated EGF-R phosphorylation is resistant to staurosporine, an extremely potent inhibitor of PKC. Analysis of tryptic 32P-phosphopeptides reveals that Thr654, the unique site of PKC-mediated phosphorylation, is not phosphorylated in cytokine-treated cells, but a different, relatively acidic, peptide containing phosphoserine can be detected instead.
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PMID:Down-modulation of epidermal growth factor receptor affinity in fibroblasts treated with interleukin 1 or tumor necrosis factor is associated with phosphorylation at a site other than threonine 654. 168 28

Staurosporine is a potent microbial inhibitor of a number of protein kinases, including protein kinase C, cyclic AMP-dependent kinase, and the tyrosine kinase pp60src. We have used staurosporine to investigate the role of phosphorylation in the regulation of the epidermal growth factor (EGF) receptor in both human epidermal carcinoma A431 cells and mouse Swiss 3T3 fibroblasts. We report here that staurosporine treatment causes enhancement in high affinity EGF binding and a decrease in the phosphorylation state of the unstimulated receptor at a number of residues, including threonine 669. Staurosporine also antagonizes the inhibition of high affinity EGF binding and the increase in phosphorylation state of the unstimulated EGF receptor by phorbol esters and the calcium ionophore A23187. Staurosporine is an effective inhibitor of the EGF-stimulated receptor tyrosine kinase in vitro and thus does not enhance EGF stimulation of EGF receptor autophosphorylation in vivo. These results suggest that phosphorylation plays a major role in the regulation of the high affinity binding state of the EGF receptor in both unstimulated and mitogenically activated cells.
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PMID:Regulation of the epidermal growth factor receptor by growth-modulating agents: effects of staurosporine, a protein kinase inhibitor. 168 32


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