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)

Previous studies show that angiotensin II (Ang II) increases phosphoinositide turnover in cultured neonatal heart cells. Ang II has also been shown to transiently increase spontaneous beating behavior in these cells. In this study we seek to identify the molecular mechanism underlying this rapid (3-5-minute) desensitization. Time-course studies on the accumulation of [3H]inositol phosphates indicate that the loss in functional responsiveness correlates with reduced efficacy of Ang II to activate the phosphoinositide path. Binding studies with 125I-Ang II revealed that there was no change in surface receptor binding capacity during the time in which desensitization developed. Normal phosphoinositide and functional responses are observed when desensitized cells are treated with probes that activate the cardiac phosphoinositide pathway at discrete steps. These studies reveal that the functional status of the major components of the phosphoinositide signaling pathway, including G proteins, phospholipase C, and protein kinase C (PKC), are normal during maintained Ang II desensitization. To study the potential role of PKC in Ang II desensitization, the cells are treated with TPA for 24 hours, which downregulates PKC activity. PKC-depleted cells rapidly desensitize after Ang II application. We conclude that the selective Ang II-evoked biochemical/functional desensitization involves inhibition at the level of the receptor, rather than at a component downstream in the path, and that this process is independent of PKC and loss of surface binding capacity.
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PMID:Angiotensin-induced desensitization of the phosphoinositide pathway in cardiac cells occurs at the level of the receptor. 165 18

We studied the effects of changing the intracellular Ca level ([Ca]i) and activating protein kinase C on the cardiac T and L Ca channels in single canine ventricular and Purkinje cells. Lowering [Ca]i increased the L current but decreased the T current, whereas elevating [Ca]i caused opposite changes. In ventricular cells, isoproterenol (1 microM) increased the amplitude of not only the L but also the T currents; the latter effect probably was secondary to a rise in [Ca]i following the augmentation of the L current. 12-O-tetradecanoylphorbol-13-acetate (TPA, 100 nM) decreased the T current but first increased and then decreased the L current. The TPA effects on the T and L currents were not mimicked by a phorbol ester that does not activate PKC (4 alpha-phorbol 12,13-didecanoate) and were prevented by a protein kinase inhibitor (H-8), confirming the involvement of PKC activity in these modulatory processes. We conclude that elevating [Ca]i and activating PKC have opposite effects on the T and L Ca currents in canine cardiac cells. The extent and time course of the changes in these two intracellular messengers will most likely determine the effects on the two cardiac Ca currents of neurotransmitters and hormones that can activate phospholipase C.
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PMID:Different effects of intracellular Ca and protein kinase C on cardiac T and L Ca currents. 165 11

The maintenance of optimal steroidogenesis in adrenocortical cells primarily depends on the chronic action of ACTH to promote the synthesis of the various steroid metabolizing enzymes. In the steroidogenic pathway, the ratio of 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) to 17 alpha-hydroxylase cytochrome P450 (P-450(17 alpha)) plays a key role in determining the final steroid products released by adrenal cells. The differences in these enzymes are particularly important when one considers the adrenal zones and the secretion of the zone-specific steroids. In the present study we have investigated the regulation of 3 beta HSD with regard to its enzyme activity, levels of protein and changes in specific mRNA encoding for this enzyme. Following eight days in primary culture, bovine adrenocortical (BAC) cells were found to respond to both ACTH and Bu2 cAMP by increased cortisol production. In addition, 3 beta HSD activity, enzyme protein and mRNA levels were increased in response to both factors. The increases varied from 2-fold for activity to 5-7 fold for mRNA. ACTH and Bu2cAMP also greatly increased P-450(17 alpha) from the near undetectable levels in control cells. In order to examine the possibility of differential regulation of these adrenal steroidogenic enzymes we determined the effects of angiotensin II (A-II) and transforming growth factor beta (TGF beta) on the levels of these enzymes. Both of these factors decreased the ACTH-stimulated levels of P-450(17 alpha) enzyme and mRNA to near nondetectable levels observed within control cells. In addition, these compounds inhibited the ACTH induction of 3 beta HSD. While the mechanism of TGF beta action is not clear, A-II probably is acting through protein kinase C. Indeed the protein kinase C activating phorbol ester, TPA, mimicked the inhibitory effects of A-II on 3 beta HSD and P450(17 alpha). It is important to point out, however, that the effects of A-II and TGF beta on P450(17 alpha) activity appeared more pronounced than their action of 3 beta HSD. This observation may relate to the relative stability of 3 beta HSD as compared to P450(17 alpha). Taken together these data indicate that, while A-II and TGF beta each decrease the levels of steroid-metabolizing enzymes, a differential regulation is observed in that P-450(17 alpha) protein and activity levels are much more sensitive to treatment with these factors.
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PMID:Regulation of 3 beta-hydroxysteroid dehydrogenase in adrenocortical cells: effects of angiotensin-II and transforming growth factor beta. 165 33

Basic fibroblast growth factor (bFGF) stimulates mitogenesis of Balb/c 3T3 fibroblast cells. This stimulation may be mediated by multiple signal pathways as it is accompanied by the formation of inositol phosphates, activation of PKC (protein kinase C) and a decrease in intracellular cAMP levels. The multiple positive and negative pathways implicated for FGF-induced mitogenesis may interact and each may contribute in varying degrees to the final cellular response. At least two types of G-proteins may be involved in the intracellular signalling pathways of FGF. Pertussis toxin blocks FGF and TPA (12-O-tetradecanoylphorbol-13-acetate) induced. PKC-mediated mitogenesis and also the associated fall in intracellular cAMP levels. However, pertussis toxin has no effect upon FGF-induced inositol phosphates formation. Thus, inhibition of mitogenesis by pertussis toxin may involve pertussis toxin sensitive G-proteins which may affect at least two separate putative signal pathways involving adenylate cyclase and protein kinase C. Pertussis toxin insensitive G-proteins may also be involved in coupling the FGF receptor to phosphoinositidase C.
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PMID:Studies on the mechanisms of signalling and inhibition by pertussis toxin of fibroblast growth factor-stimulated mitogenesis in Balb/c 3T3 cells. 165 71

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

We obtained a Ca(2+)-independent but 12-O-tetradecanoyl phorbol ester (TPA).phospholipid-activated protein kinase from rat embryo fibroblast 3Y1 cells by succeeding steps of DEAE-cellulose, H-9 affinity, and hydroxylapatite chromatography. This kinase was separated chromatography. This kinase was separated from a conventional PKC (Type III), by H-9 affinity column chromatography. The major peak from H-9 affinity column was eluted at 0.4 M of arginine and on the following step of hydroxylapatite column chromatography, at the KPO4 concentration of 0.1 M. The enzyme could be stimulated by phospholipids and by the tumor promoter TPA, but did not respond to calcium. The Ca(2+)-independent, phospholipid-activated protein kinase activity was susceptible to the protein kinase C inhibitors H-7 and K252a, but showed a phospholipid dependency and substrate specificity distinct from the conventional types of PKC. This protein kinase did not react with monoclonal antibodies against Types I, II, and III PKC. The activity of this enzyme was specifically reduced by immunoprecipitation, depending on the concentration of the polyclonal antibody, PC-delta, which was raised against a peptide synthesized according to a sequence of rat brain nPKC delta. The enzyme had a Mr of 76,000 as estimated by Western blotting. These results provide evidence for a unique type of Ca(2+)-independent, phospholipid-activated kinase, as expressed in 3Y1 cells.
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PMID:Ca(2+)-independent, phospholipid-activated protein kinase in 3Y1 cells. 165 33

We have previously shown that after peripheral nerve lesion the synthesis of NGF is induced in cells of the nerve sheath (Heumann et al., 1987a). Further analysis led to the identification of growth factors and intracellular mechanisms responsible for this induction in sciatic fibroblasts (Lindholm et al., 1988; Hengerer et al., 1990). The present work aimed at the elucidation of the regulation of NGF synthesis in Schwann cells. A variety of cytokines and peptide growth factors, including interleukin-1 (IL-1) and platelet-derived growth factor (PDGF), which are known to increase NGF-mRNA in fibroblasts and astrocytes, failed to do so in Schwann cell cultures. Forskolin (FK), an activator of adenylate cyclase, increased the level of NGF-mRNA eightfold within 3 hr of incubation. The effect of FK on NGF-mRNA was mimicked by analogs of cAMP but not by dideoxyforskolin, an FK derivative not activating adenylate cyclase. Application of norepinephrine and isoproterenol also augmented the NGF-mRNA content. Pretreatment of Schwann cells with N-[2-(methylamino)ethyl]-5-isoquinoline sulfonamide dihydrochloride (H-8), an inhibitor of cyclic-nucleotide-dependent protein kinases, decreased both basal and elevated levels of NGF-mRNA. Ionomycin, a Ca2+ ionophore, and phorbol 12-myristate 13-acetate (TPA), an activator of protein kinase C, potentiated the effect of FK in an H-8-sensitive manner. We show that the action of FK is independent of changes in mRNA stability and of protein synthesis. Thus, in cultured Schwann cells upregulation of NGF-mRNA expression seems to be mainly achieved by a cAMP-triggered transcriptional activation of the NGF gene. Another striking difference between various glial cell types was revealed by application of transforming growth factor beta-1 (TGF-beta 1), which is the strongest inducer of NGF-mRNA in cultured astrocytes (Lindholm et al., 1990). Schwann cells responded to TGF-beta 1 by decreasing basal as well as FK-induced NGF-mRNA levels. Together with previously published work, our results show that cell-type-specific mechanisms not only account for the different control of NGF expression in neurons as compared to glial cells, but also reveal a surprising specificity of regulatory mechanisms in different non-neuronal cell types, even those derived from the same tissue such as fibroblasts and Schwann cells of peripheral nerves.
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PMID:Cell-type-specific regulation of nerve growth factor (NGF) synthesis in non-neuronal cells: comparison of Schwann cells with other cell types. 165 45

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

Recent evidence has been presented that follicle-stimulating hormone (FSH) stimulates the induction of granulosa cell c-fos protooncogene mRNA in vivo (Pennybacker and Herman (1989) J. Cell Biol. 109, 151A; Delidow et al. (1990) Endocrinology 126, 2302-2306), yet the mechanisms by which FSH induces c-fos mRNA expression have not been delineated. To elucidate the mechanisms of FSH-dependent c-fos mRNA expression, we measured the time and dose dependence of c-fos mRNA levels using Northern blot analysis in intact ovaries and cultured granulosa cells in response to FSH. In intact ovaries, FSH-induced c-fos mRNA expression was time dependent with maximal expression at 90 min post FSH injection, while in cultures of granulosa cells obtained from estrogen-primed immature female rats, c-fos mRNA levels were highest after 30 min exposure to FSH and at a concentration of 100 ng/ml. Neither 8-bromo adenosine 3',5'-cyclic monophosphate (8-br-cAMP), at doses ranging from 0.1 to 10 mM, nor 100 microM forskolin (in the presence or absence of 200 microM isobutyl-methylxanthine) or luteinizing hormone (LH, 100 ng/ml) were able to mimic FSH-induced c-fos mRNA expression in granulosa cell cultures. However, tetradecanoyl-13-phorbol acetate (TPA, 200 nM) was able to induce c-fos mRNA expression. The protein kinase C (PKC) inhibitors H-7 (0.3-30 microM) and staurosporine (0.75 micrograms/ml) blocked FSH-induced c-fos mRNA expression in cultured granulosa cells while HA 1004, an inhibitor of cGMP- and cAMP-dependent protein kinases at 30 microM had no effect on TPA-induced c-fos expression, and only minimally inhibited FSH-induced c-fos expression. Both FSH (100 ng/ml) and forskolin (3 microM) increased progesterone production in cultured granulosa cells. These data support the hypothesis that FSH specifically induces c-fos mRNA expression by a PKC-dependent mechanism and that the cAMP arm of the FSH response pathway is operant in these cells.
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PMID:Follicle-stimulating hormone increases c-fos mRNA levels in rat granulosa cells via a protein kinase C-dependent mechanism. 165 43

The mechanisms of muscarinic receptor-linked increase in cAMP accumulation in SH-SY5Y human neuroblastoma cells has been investigated. The dose-response relations of carbachol-induced cAMP synthesis and carbachol-induced rise in intracellular free Ca2+ were similar. The stimulated cAMP synthesis was inhibited by about 50% when cells were entrapped with the Ca2+ chelator BAPTA or in the presence of the protein kinase C (PKC) inhibitor staurosporine. Production of cAMP could be induced also by the Ca2+ ionophore, ionomycin and by TPA, an activator of PKC. When added together TPA and ionomycin had a synergistic effect. When cAMP synthesis was activated with cholera toxin, PGE1 or PGE1 + pertussis toxin carbachol stimulated cAMP production to the same extent as in control cells. Ca2+ and protein kinase C thus seem to be the mediators of muscarinic-receptor linked cAMP synthesis by a direct action on adenylate cyclase.
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PMID:Muscarinic receptor-linked elevation of cAMP in SH-SY5Y neuroblastoma cells is mediated by Ca2+ and protein kinase C. 165 8


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