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 extracellular Ca2+ (Ca2+(o))-sensing receptor (CaR) is a G protein-coupled receptor that activates phospholipase C (PLC). In the present studies, we assessed Ca2+(o)-dependent changes in the generation of inositol phosphates (IP), free arachidonic acid (AA), and phosphatidylbutanol (PtdBtOH) by PLC, phospholipase A2 (PLA2), and phospholipase D (PLD), respectively, in bovine parathyroid cells as well as in wild-type or CaR-transfected human embryonic kidney (HEK293) cells (HEK-WT and HEK-CaR, respectively). Elevated Ca2+(o) increased the formation of IPs in parathyroid cells as well in HEK-CaR but not in HEK-WT cells. High Ca2+(o) also elicited time- and dose-dependent increases in PtdBtOH in parathyroid cells and HEK-CaR but not in HEK-WT cells. Brief treatment of parathyroid and HEK-CaR cells with an activator of protein kinase C (PKC), phorbol 12-myristate,13-acetate (PMA), stimulated PLD activity at both low and high Ca2+(o). Moreover, high Ca2+(o)-stimulated PLD activity was abolished following down-regulation of PKC by overnight phorbol myristate acetate (PMA) pretreatment, suggesting that CaR-mediated activation of PLD depends largely upon stimulation of PKC. High Ca2+(o) likewise increased the release of free AA in parathyroid and HEK-CaR but not in HEK-WT cells. Mepacrine, a general PLA2 inhibitor, and AACOCF3, an inhibitor of cytosolic PLA2, reduced AA release in parathyroid cells at high Ca2+(o), suggesting a major role for PLA2 in high Ca2+(o)-elicited AA release. Pretreatment of parathyroid cells with PMA stimulated release of AA at low and high Ca2+(o), while a PKC inhibitor, chelerythrine, reduced AA release at high Ca2+(o) to the level observed with low Ca2+(o) alone. Thus, PKC contributes importantly to the high Ca2+(o)-evoked, CaR-mediated activation of not only PLD but also PLA2. Finally, high Ca2+(o)-stimulated production of IP, PtdBtOH, and AA all decreased substantially in parathyroid cells cultured for 4 days, in which expression of the CaR decreases by 80% or more, consistent with mediation of these effects by the receptor. Thus, the CaR activates, directly or indirectly, at least three phospholipases in bovine parathyroid and CaR-transfected HEK293 cells, providing for coordinate, receptor-mediated regulation of multiple signal transduction pathways in parathyroid and presumably other CaR-expressing cells.
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PMID:The Ca2+-sensing receptor (CaR) activates phospholipases C, A2, and D in bovine parathyroid and CaR-transfected, human embryonic kidney (HEK293) cells. 914 37

Many receptors that couple to heterotrimeric guanine-nucleotide binding proteins (G proteins) have been shown to mediate rapid activation of the mitogen-activated protein kinases Erk1 and Erk2. In different cell types, the signaling pathways employed appear to be a function of the available repertoire of receptors, G proteins, and effectors. In HEK-293 cells, stimulation of either alpha1B- or alpha2A-adrenergic receptors (ARs) leads to rapid 5-10-fold increases in Erk1/2 phosphorylation. Phosphorylation of Erk1/2 in response to stimulation of the alpha2A-AR is effectively attenuated by pretreatment with pertussis toxin or by coexpression of a Gbetagamma subunit complex sequestrant peptide (betaARK1ct) and dominant-negative mutants of Ras (N17-Ras), mSOS1 (SOS-Pro), and Raf (DeltaN-Raf). Erk1/2 phosphorylation in response to alpha1B-AR stimulation is also attenuated by coexpression of N17-Ras, SOS-Pro, or DeltaN-Raf, but not by coexpression of betaARK1ct or by pretreatment with pertussis toxin. The alpha1B- and alpha2A-AR signals are both blocked by phospholipase C inhibition, intracellular Ca2+ chelation, and inhibitors of protein-tyrosine kinases. Overexpression of a dominant-negative mutant of c-Src or of the negative regulator of c-Src function, Csk, results in attenuation of the alpha1B-AR- and alpha2A-AR-mediated Erk1/2 signals. Chemical inhibitors of calmodulin, but not of PKC, and overexpression of a dominant-negative mutant of the protein-tyrosine kinase Pyk2 also attenuate mitogen-activated protein kinase phosphorylation after both alpha1B- and alpha2A-AR stimulation. Erk1/2 activation, then, proceeds via a common Ras-, calcium-, and tyrosine kinase-dependent pathway for both Gi- and Gq/11-coupled receptors. These results indicate that in HEK-293 cells, the Gbetagamma subunit-mediated alpha2A-AR- and the Galphaq/11-mediated alpha1B-AR-coupled Erk1/2 activation pathways converge at the level of phospholipase C. These data suggest that calcium-calmodulin plays a central role in the calcium-dependent regulation of tyrosine phosphorylation by G protein-coupled receptors in some systems.
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PMID:Ras-dependent mitogen-activated protein kinase activation by G protein-coupled receptors. Convergence of Gi- and Gq-mediated pathways on calcium/calmodulin, Pyk2, and Src kinase. 923 1

Downregulation of insulin receptor tyrosine kinase (IRK) activity yields to impaired insulin signalling and contributes to the pathogenesis of cellular insulin resistance. Activation of protein kinase C (PKC) by different agents is associated with an inhibition of IRK activity in various cell types. There is evidence that this effect on IRK activity might be mediated through phosphorylation of specific serine residues of the insulin receptor beta-subunit. Neither the domains of the IRK where inhibiting serine phosphorylation occurs nor the PKC isoform responsible for IRK inhibition have been identified. PKC consists of a family of at least 12 isoforms. The aim of the present study was to determine which PKC isoform might be capable of IRK inhibition. The human insulin receptor and the PKC isoforms alpha, beta 1, beta 2, gamma, delta, epsilon, eta, theta and zeta were overexpressed in human embryo kidney fibroblasts (HEK 293 cells) in order to answer this question. PKCs were activated by preincubation with the phorbolester (TPA) (10(-7) mol/l) following insulin stimulation of the cells. When the IRK was coexpressed with the PKC isoforms beta 1 and beta 2, a 50 +/- 15.7 and 45 +/- 10.1% inhibition of tyrosine autophosphorylation of IRK was observed while coexpression with the other isoforms did not significantly modify IRK autophosphorylation. The data suggest that the PKC isoforms beta 1 and beta 2 might be candidates for insulin receptor inhibition.
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PMID:Protein kinase C isoforms beta 1 and beta 2 inhibit the tyrosine kinase activity of the insulin receptor. 924 10

Adenylyl cyclases of the type II family differ from other subforms in that they are conditionally stimulated via alpha(s)/betagamma subunits and regulated by PKC mediated phosphorylation. AC II, stably expressed in HEK 239 cells, was incubated with the PKC activator tetradecanoylphorbol acetate (TPA). Using cells metabolically labeled with [32P]phosphate, TPA caused concerted stimulation of basal and forskolin activated adenylyl cyclase together with incorporation of [32P]phosphate into AC II protein. Enhanced phosphorylation was also indicated by a monoclonal anti-phosphothreonine antibody. Assignment of TPA-induced [32P]phosphate-incorporation to specific sites was achieved by a combination of chemical and immunochemical methods. Three out of five [32P]labeled peptides that were generated by fragmentation with N-chlorosuccinimide were also recognized by the monoclonal antibody BBC-4 [S. Mollner, T. Pfeuffer, Eur. J. Biochem. 171 (1988) 265-271] directed against an epitope 8 kDa from the extreme C-terminus. These findings suggested Ser-871 (consensus sequence ARSLK) and Thr-1057 (CTCR) as acceptor candidates of phorbolester induced phosphoryl transfer.
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PMID:Adenylyl cyclase type II is stimulated by PKC via C-terminal phosphorylation. 936 62

Ca2+/CaM-dependent protein kinase II (CaM-KII) can phosphorylate and potentiate responses of alpha-amino3-hydroxyl-5-methyl-4-isoxazole-propionate-type glutamate receptors in a number of systems, and recent studies implicate this mechanism in long term potentiation, a cellular model of learning and memory. In this study we have identified this CaM-KII regulatory site using deletion and site-specific mutants of glutamate receptor 1 (GluR1). Only mutations affecting Ser831 altered the 32P peptide maps of GluR1 from HEK-293 cells co-expressing an activated CaM-KII. Likewise, when CaM-KII was infused into cells expressing GluR1, the Ser831 to Ala mutant failed to show potentiation of the GluR1 current. The Ser831 site is specific to GluR1, and CaM-KII did not phosphorylate or potentiate current in cells expressing GluR2, emphasizing the importance of the GluR1 subunit in this regulatory mechanism. Because Ser831 has previously been identified as a protein kinase C phosphorylation site (Roche, K. W., O'Brien, R. J., Mammen, A. L., Bernhardt, J., and Huganir, R. L. (1996) Neuron 16, 1179-1188), this raises the possibility of synergistic interactions between CaM-KII and protein kinase C in regulating synaptic plasticity.
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PMID:Identification of the Ca2+/calmodulin-dependent protein kinase II regulatory phosphorylation site in the alpha-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate-type glutamate receptor. 940 43

The effect of nicotine on the major human neuronal nicotinic receptor (alpha 4 beta 2 subtype) was studied in permanently transfected HEK 293 cells. Prolonged exposure to low concentrations of nicotine (1 microM) increased epibatidine binding but functionally deactivated the nicotinic receptor, abolishing Ca2+ influx in response to an acute nicotine challenge. Deactivation could also be caused by down-regulating protein kinase C (PKC) activity with 0.5 microM phorbol-12,13-dibutyrate or briefly incubating cells with the PKC inhibitor NPC-15437. Recovery from receptor deactivation caused by either nicotine treatment or PKC inhibition occurred slowly (4-6 hr). Reversal of nicotine-induced deactivation was accelerated by the addition of inhibitors of protein phosphatases 2A and 2B. These data suggest a hypothetical mechanism of nicotine-induced deactivation that involves dephosphorylation of nicotinic receptors at PKC phosphorylation sites.
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PMID:Functional deactivation of the major neuronal nicotinic receptor caused by nicotine and a protein kinase C-dependent mechanism. 941 21

We examined whether histamine could regulate cell proliferation and expression of the early response gene c-fos in HEK-293 cells stably transfected with the human H2 receptor (HEK-H2). Histamine stimulated [3H]thymidine incorporation [50% effective concentration (EC50) = 3.6 x 10(-6) M] in HEK-H2 cells in a cimetidine-sensitive manner and increased c-fos mRNA in a time-dependent fashion, reaching maximal induction after 30 min. Histamine induced luciferase activity in HEK-H2 cells transiently transfected with a construct containing the luciferase reporter gene (Luc) coupled to the serum response element (SRE) of the c-fos gene promoter (EC50 = 1.5 x 10(-6) M) or a plasmid containing the SRE core fragment (bases -320 to -298). The protein kinase C (PKC) inhibitor staurosporine and long-term pretreatment of HEK cells with phorbol ester inhibited the effect of histamine on PKC activation, SRE-Luc activity, and [3H]thymidine incorporation. We have demonstrated that activation of the human H2 receptor can lead to induction of c-fos gene transcription and cell proliferation through a PKC-dependent mechanism.
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PMID:Activation of the human histamine H2 receptor is linked to cell proliferation and c-fos gene transcription. 943 11

Antidepressant-sensitive serotonin (5-hydroxytryptamine, 5HT) transporters (SERTs) are responsible for efficient synaptic clearance of extracellular 5HT. Previously (Qian, Y., Galli, A., Ramamoorthy, S., Risso, S., DeFelice, L. J., and Blakely, R. D. (1997) J. Neurosci. 17, 45-47), we demonstrated that protein kinase (PKC)-linked pathways in transfected HEK-293 cells lead to the internalization of cell-surface human (h) SERT protein and a reduction in 5HT uptake capacity. In the present study, we report that PKC activators rapidly, and in a concentration-dependent manner, elevate the basal level of hSERT phosphorylation 5-6-fold. Similarly, protein phosphatase (PP1/PP2A) inhibitors down-regulate 5HT transport and significantly elevate hSERT 32P incorporation, effects that are additive with those of PKC activators. Moreover, hSERT phosphorylation induced by beta-phorbol 12-myristate 13-acetate is abolished selectively by the PKC inhibitors staurosporine and bisindolylmaleimide I, whereas hSERT phosphorylation induced by phosphatase inhibitors is insensitive to these agents at comparable concentrations. Protein kinase A and protein kinase G activators fail to acutely down-regulate 5HT uptake but significantly enhance hSERT phosphorylation. Basal hSERT and okadaic acid-induced phosphorylation were insensitive to chelation of intracellular calcium and Ca2+/calmodulin-dependent protein kinase inhibitors. Together these results reveal hSERT to be a phosphoprotein whose phosphorylation state is likely to be tightly controlled by multiple kinase and phosphatase pathways that may also influence the transporter's regulated trafficking.
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PMID:Phosphorylation and regulation of antidepressant-sensitive serotonin transporters. 944 97

Ceramides are reported to stimulate different effector systems, among them atypical protein kinases C (PKCs). When HEK 293 cells, stably expressing adenylyl cyclase type II (AC II), were treated with various ceramide derivatives, adenylyl cyclase activity was enhanced 8-15-fold. The stimulation by the most potent analog, C18/C24 ceramide, was comparable to that by the phorbolester TPA. The stimulatory effect of ceramide was not restricted to AC II, although the type I and type V enzymes were affected less dramatically. Unexpectedly, the dihydro derivatives of ceramides, generally serving as non-activating controls, exhibited only slightly lower stimulation than ceramides, whereas short-chain ceramides (e.g. C2) were without effect. The action of ceramides was at least partially inhibited by okadaic acid, suggesting involvement of a phosphatase. Furthermore, ceramides and TPA operated synergistically. While the PKC inhibitor staurosporine counteracted the action of phorbol-esters, it significantly (2.5x) enhanced the effect of ceramides.
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PMID:Differential effects of ceramides upon adenylyl cyclase subtypes. 949 8

Protein kinases and phosphatases are targeted through association with anchoring proteins that tether the enzymes to subcellular structures and organelles. Through in situ fluorescent techniques using a Green Fluorescent Protein tag, we have mapped membrane-targeting domains on AKAP79, a multivalent anchoring protein that binds the cAMP-dependent protein kinase (PKA), protein kinase C (PKC) and protein phosphatase 2B, calcineurin (CaN). Three linear sequences termed region A (residues 31-52), region B (residues 76-101) and region C (residues 116-145) mediate targeting of AKAP79 in HEK-293 cells and cortical neurons. Analysis of these targeting sequences suggests that they contain putative phosphorylation sites for PKA and PKC and are rich in basic and hydrophobic amino acids similar to a class of membrane-targeting domains which bind acidic phospholipids and calmodulin. Accordingly, the AKAP79 basic regions mediate binding to membrane vesicles containing acidic phospholipids including phosphatidylinositol-4, 5-bisphosphate [PtdIns(4,5)P2] and this binding is regulated by phosphorylation and calcium-calmodulin. Finally, AKAP79 was shown to be phosphorylated in HEK-293 cells following stimulation of PKA and PKC, and activation of PKC or calmodulin was shown to release AKAP79 from membrane particulate fractions. These findings suggest that AKAP79 might function in cells not only as an anchoring protein but also as a substrate and effector for the anchored kinases and phosphatases.
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PMID:Membrane-targeting sequences on AKAP79 bind phosphatidylinositol-4, 5-bisphosphate. 954 38


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