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 commitment of myogenically determined cells to terminal differentiation can be modulated by a variety of agents, including growth factors and activated oncogenes. We have examined the effect of interleukin 1 alpha (IL-1 alpha) on the terminal differentiation of a normal myogenically determined cell line and two myogenically determined, differentiation competent cell lines which contain either one or six copies of the activated c-Ha-ras oncogene. Treatment of all cell lines with IL-1 alpha decreased but did not totally inhibit terminal myogenic differentiation. Over the range of IL-1 alpha concentrations assayed (1-40 ng/ml), the c-Ha-ras transformed cell lines demonstrated a significantly greater sensitivity to the inhibitory effects of IL-1 alpha. The inhibition of differentiation was not the result of enhanced proliferation. Interestingly, transformation with activated c-Ha-ras resulted in a decrease in IL-1 alpha receptor number and affinity. The enhanced IL-1 alpha responsiveness of the ras transformants was not the result of increased proliferation or changes in either ras gene expression or protein kinase C activity. IL-1 alpha treatment decreased the steady-state levels of both MyoD1 and myogenin transcripts in the c-Ha-ras transformed but not the normal myogenic cell line. Further studies are required to determine the mechanism(s) responsible for the increased sensitivity of the c-Ha-ras transformed cultures to the inhibitory effects of IL-1 alpha.
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PMID:Interleukin 1 alpha mediated inhibition of myogenic terminal differentiation: increased sensitivity of Ha-ras transformed cultures. 132 82

Myogenin belongs to a family of myogenic helix-loop-helix (HLH) proteins that activate muscle transcription through binding to a conserved DNA sequence associated with numerous muscle-specific genes. Fibroblast growth factor (FGF) inhibits myogenesis by inactivating myogenic HLH proteins. We show that activated protein kinase C (PKC) can substitute for FGF and inhibit transcriptional activity of myogenic HLH proteins. In transfected cells, FGF induces phosphorylation of a conserved site in the DNA-binding domain of myogenin. This site is phosphorylated by PKC in vivo and in vitro and mediates repression of the myogenic program through a loss in DNA binding activity. A myogenin mutant lacking the PKC phosphorylation site is not repressed by FGF, confirming this site as a molecular target for FGF-dependent repression of muscle transcription. These results establish a direct link between the signal transduction pathways that inhibit myogenesis and the transcription factors directly activating muscle-specific genes.
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PMID:FGF inactivates myogenic helix-loop-helix proteins through phosphorylation of a conserved protein kinase C site in their DNA-binding domains. 133 66

Protein kinase C has previously been implicated in the regulation of chicken acetylcholine receptor (AChR) gene expression. To investigate the molecular basis of this regulation, the promoter of the AChR alpha-subunit (alpha AChR) gene was linked to a reporter gene and introduced into cultured chick myotubes by transient transfection. Treatment of myotubes with protein-kinase-C-activating phorbol esters was found to inhibit promoter activity. These inhibitory actions were mediated by promoter sequences between nucleotides -110 and -45, relative to the start point of transcription of the alpha AChR gene. In particular, phorbol-ester responsiveness could be conferred by a short DNA sequence that contains one of the two MyoD binding sites of the alpha AChR gene muscle-specific enhancer. 12-O-Tetradecanoylphorbol 13-acetate was found to inhibit rapidly and potently the expression of mRNAs coding for the myogenic regulators CMD1 and myogenin. Moreover, its inhibitory effect on the alpha AChR gene promoter could be attenuated by cotransfection of a MyoD1 expression vector. These results provide a molecular basis for the previously demonstrated involvement of protein kinase C in the regulation of alpha AChR biosynthesis. In addition, they lend further support to the notion that myogenic proteins play an important role in the control of alpha AChR gene expression.
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PMID:Phorbol esters inhibit the activity of the chicken acetylcholine receptor alpha-subunit gene promoter. Role of myogenic regulators. 176 95

Human rhabdomyosarcoma RD cells express the myogenic regulatory factors MyoD and myogenin but differentiate spontaneously very poorly. Prolonged treatment of RD cells with the protein kinase C (PKC) activator 12-O-tetradecanoylphorbol-13-acetate (TPA) induces growth arrest and myogenic differentiation as shown by the accumulation of alpha-actin and myosin light and heavy chains, without affecting the expression of MyoD and myogenin. In this study, we show that short-term phorbol ester treatment of the cultures is sufficient to trigger myogenic differentiation but not growth arrest. Furthermore, PKC inhibitors, such as staurosporine or calphostin C, prevent TPA-induced differentiation but not cell growth arrest. These data suggest that the two events are mediated by different pathways; a possible interpretation is that the activation of one or more PKC isoforms mediates the induction of differentiation, whereas the down-regulation of the same or different isoforms mediates the growth arrest. To address the mechanism whereby TPA affects cell growth and differentiation in RD cells, we first analyzed PKC isoenzyme distribution. We found that RD cells express the alpha, beta 1, gamma, and sigma PKC isoenzymes. Only the alpha isoform is exclusively found in the soluble fraction, but it translocates to the membrane fraction within 5 min of TPA treatment and is completely down-regulated after 6 h. The other isoenzymes are found associated to both the soluble and the particulate fractions and are down-regulated after long-term TPA treatment. By immunofluorescence analysis, we show that the PKC alpha down-regulation is specific for those cells that respond to TPA by activating the muscle phenotype. We propose that TPA-induced differentiation in RD cells is mediated by the transient activation of PKC alpha, which activates some of the intracellular events that are necessary for MyoD and myogenin transacting activity and for the induction of terminal differentiation of RD cells. By contrast, the constitutively active beta 1 and sigma are responsible for the maintenance of cell growth, and their down-regulation is responsible for long-term TPA-induced cell growth arrest.
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PMID:Rapid activation and down-regulation of protein kinase C alpha in 12-O-Tetradecanoylphorbol-13-acetate-induced differentiation of human rhabdomyosarcoma cells. 754 6

We have carried out an analysis of 833 bp of the chick myogenin gene 5' flanking sequence. A 131-bp segment of this upstream region, which contains a CANNTG promoter element (E box) and a myocyte-enhancer-binding-factor-2 (MEF-2) site, acts as a full promoter. It resembles the human and the mouse myogenin promoters in the structure and disposition of regulatory elements, including a TATA box and the transcription start site. Examination of eight myogenic factor/E protein combinations cotransfected with several myogenin promoter constructs into HeLa cells, reveals that the chick myogenin/E2-5 combination maximally activates the myogenin promoter. This activation is mediated through the E box motif; the MEF-2 site limits the factor combinations that can activate the myogenin promoter and enhances activation by myogenin/E2-5. We have found previously that activation of protein kinase C inactivates the transcription of the chick myogenin gene [Huang, C.-F., Neville, C. M. & Schmidt, J. (1993) Control of myogenic factor genes by the membrane depolarization/protein kinase C cascade in chick skeletal muscle, FEBS Lett. 319, 21-25]. In this study, we show that the activation of protein kinase C inhibits the factor bound to the E box, which thereupon negatively regulates the activity of the MEF-2 binding protein.
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PMID:The role of the CANNTG promoter element (E box) and the myocyte-enhancer-binding-factor-2 (MEF-2) site in the transcriptional regulation of the chick myogenin gene. 760 Nov 28

We have analyzed the potential role of myogenin in the regulation by electrical activity of the expression of the acetylcholine receptor (AChR) alpha-subunit gene in cultured chick embryonic myotubes. The state of phosphorylation of myogenin was followed by 32P-labeling and immunoprecipitation with an anti-myogenin antibody. In electrically active myotubes myogenin is phosphorylated, while it is dephosphorylated in electrically silent myotubes following tetrodotoxin (TTX) treatment. Accordingly, nuclear protein kinase C (PKC) activity decreases in TTX-treated myotubes. Myogenin dephosphorylation is also observed upon incubation of myotubes with GF109203X, a pharmacological agent which specifically inhibits PKC activity. Both treatments cause similar increases in the expression of the AChR protein. The effects are not additive. Thus TTX and GF109203X most probably affect a common process. Recombinant chick myogenin binds to myogenic sites (E boxes) present in the AChR alpha-subunit promoter but loses this binding capacity after phosphorylation. As a working hypothesis we propose that repression of AChR biosynthesis by electrical activity results, at least partly, from phosphorylation of myogenin via the PKC pathway.
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PMID:Phosphorylation of myogenin in chick myotubes: regulation by electrical activity and by protein kinase C. Implications for acetylcholine receptor gene expression. 811 18

MRF4 is a member of the muscle-specific basic helix-loop-helix transcription factor family that also includes MyoD, myogenin, and Myf-5. Each of these proteins, when overexpressed in fibroblasts, converts the cells to differentiated muscle fibers that express several skeletal muscle genes, such as those for alpha-actin, muscle creatine kinase, and troponin I. Despite the fact that MRF4 functions as a positive transcriptional regulator, the MRF4 protein is subject to negative regulation by a variety of agents, most notably by exposure of cells to purified growth factors, such as basic fibroblast growth factor (bFGF). In an effort to establish whether bFGF inhibits MRF4 activity through specific posttranslational modifications, we examined whether MRF4 exists in vivo as a phosphoprotein and whether the phosphorylation status of the protein regulates its activity. Our results indicate that MRF4 is phosphorylated predominantly on serine residues, with weak phosphorylation occurring on threonine residues. Both cyclic AMP-dependent protein kinase (PKA) and protein kinase C (PKC) phosphorylate MRF4 in vitro as well as in vivo, and the overexpression of each kinase inhibits MRF4 activity and thus blocks terminal differentiation. PKC-directed phosphorylation of a conserved threonine residue (T-99) situated within the DNA-binding domain inhibits MRF4 from binding in vitro to specific DNA targets. However, although T-99 itself is essential for myogenic activity, our studies demonstrate that the phosphorylation status of T-99 does not play a major role in regulating MRF4 activity in vivo, since PKA, PKC, and bFGF inhibit the activity of MRF4 proteins in which the identified PKA and PKC sites have been mutated. We suggest that the negative regulation of MRF4 imposed by bFGF does not involve a direct modification of the protein at the identified PKA and PKC sites but instead may involve the modification of specific coregulators that interact with this muscle regulatory factor.
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PMID:Fibroblast growth factor inhibits MRF4 activity independently of the phosphorylation status of a conserved threonine residue within the DNA-binding domain. 841 99

We analysed the signaling pathways involved in myogenic differentiation of primary cultures of rat satellite cells using substances targeting the protein kinase C (PKC) and the cAMP protein kinase (PKA) pathways. We have previously shown that iso-H7, which putatively inhibits both PKC and PKA, strongly stimulates satellite cell differentiation, as well as the PKA inhibitor HA1004. In the study reported here, the effects of iso-H7 on satellite cell differentiation were compared to those observed in the presence of agents which reduce PKC activity. It was shown that treatments with the highly specific PKC inhibitor GF109203X or with 12-O-tetradecanoylphorbol 13-acetate (TPA) which induced a partial PKC downregulation, did not significantly alter myogenic differentiation. Northern blot analyses showed that iso-H7 activated the expression of myogenin but not that of MyoD mRNA. Concurrently, iso-H7 increased myosin light-chain mRNA expression. In contrast, TPA had no effect on these syntheses. Taken together, these results showed that iso-H7 did not act intracellularly as a PKC inhibitor but rather as a PKA inhibitor as previously suggested. Our results are compatible with the hypothesis that a reduction in PKA activity controls satellite cell myogenesis through an increased myogenin mRNA expression.
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PMID:The kinase inhibitor iso-H7 stimulates rat satellite cell differentiation through a non-protein kinase C pathway by increasing myogenin expression level. 881 63

Committed skeletal muscle myoblasts undergo terminal differentiation when shifted from a high-mitogen medium to a low-mitogen medium. However, expression of the myogenic regulatory factor MyoD seems to be similar in proliferating and differentiating cells, suggesting that its function is attenuated in proliferating myoblasts. To further understand the potential mechanisms that may attenuate MyoD function, we have examined the effect of posttranslational modification. By analogy with myogenin, we have examined the role of phosphorylation in regulating the function of MyoD. MyoD contains two putative protein kinase C (PKC) phosphorylation sites (Thr115 and Ser200). The former site is analogous to Thr85 within the highly conserved basic domain of myogenin that has been demonstrated to negatively regulate the myogenic differentiation functions of myogenin. To test whether hyperphosphorylation of the same PKC site in MyoD would attenuate its function, we generated a mutant MyoD with a single amino acid substitution (Thr115-Ala) that disrupts the PKC phosphorylation site (Thr115) within the conserved basic domain. Wild-type and mutant MyoD were introduced into cells using an E1, E3-deleted adenoviral vector. In mouse C3H10T1/2 fibroblasts, both wild-type and mutant MyoD induced terminal myogenic differentiation when growth factors were withdrawn from the cell culture. Consistent with these results, nuclear extracts from infected cells, but not those from uninfected cells, demonstrated complex formation with an oligonucleotide containing an E-box consensus sequence. Growth arrest was associated with the up-regulation of p21cip1, cell fusion to multinucleated myotubes, and the expression of a muscle differentiation marker (myosin heavy chain). On the other hand, when infected cells were maintained under high mitogenic conditions (in the presence of 10% fetal bovine serum), the expression of wild-type or mutant MyoD slowed cell growth and induced p21cip1. Only mutant MyoD caused cell fusion, myosin heavy chain expression, and altered mobility of the E-box oligonucleotide in gel shift assays. Furthermore, after infection, MyoD was phosphorylated, and phosphothreonine was detected in wild-type MyoD immunoprecipitated only from C3H10T1/2 cells grown under high mitogenic conditions. These results suggest that Thr115 may play an important role in the regulation of MyoD function under conditions of high mitogenesis. MyoD was also phosphorylated in malignant rhabdomyosarcoma (RMS) cells in which MyoD function was attenuated. Phosphothreonine was also detected in MyoD immunoprecipitates. Rh30 alveolar RMS cells were infected with an adenovirus expressing either wild-type or mutant MyoD. In contrast to the results in fibroblasts, when overexpressed in malignant Rh30 RMS cells, mutant MyoD arrested cell growth without inducing p21cip1 and caused cell fusion. However, no muscle differentiation markers were detected, indicating that an overexpression of mutant MyoD lacking Thr115 caused Rh30 cells to become quiescent and recapitulate at least some aspects of myogenesis (cell fusion).
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PMID:Mutation of Thr115 in MyoD positively regulates function in murine fibroblasts and human rhabdomyosarcoma cells. 975 Nov 14

The innervation-induced down-regulation of fetal-type acetylcholine receptor (AChR) expression in developing muscle fibers has largely been attributed to nerve-evoked muscle activity; however, there is increasing evidence that a neural trophic factor also contributes to this receptor down-regulation. Previous studies from this laboratory have shown that neural extracts contain a factor which decreases fetal-type AChR expression in skeletal muscle cell lines and therefore may account for the proposed inhibitory neurotrophic influence. The current study investigated possible intracellular signaling molecules involved in this receptor down-regulation and demonstrated that activation of protein kinase C and p70(S6k) appeared to be important in receptor down-regulation. Decreases in AChR density were independent of myogenin. In addition, the receptor down-regulation was independent of neuregulin, which also induces p70(S6k) activity. These studies demonstrate that neural extracts contain an inhibitory factor which can down-regulate fetal-type AChR expression independently of nerve-evoked muscle activity through intracellular signaling molecules which are known to regulate AChR expression.
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PMID:Intracellular signaling molecules involved in an inhibitory factor-induced decrease in fetal-type AChR expression. 1064 Mar 26


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