Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.12.2 (MEK)
 18,161 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Prostaglandin synthase 2 (PGS2) is an immediate-early gene induced in a variety of cellular contexts. We investigate here the transcriptional activation of the murine PGS2 gene in NIH 3T3 cells, in response to the mitogens platelet-derived growth factor (PDGF) or serum. Site-directed mutagenesis experiments demonstrate that a consensus cyclic AMP response element (CRE) in the murine PGS2 promoter is essential for optimal PGS2 gene expression in response to PDGF or to serum. Overexpression of c-Jun potentiates PDGF- or serum-induced luciferase expression from a reporter construct containing the first 371 nucleotides of the PGS2 promoter. In contrast, overexpression of other transcription factors binding to the CRE element of the PGS2 gene inhibits induction by PDGF or serum. Moreover, positioning the c-Jun activation domain next to the minimal PGS2 promoter via a GAL4 DNA binding site rather than the CRE is sufficient to permit serum or PDGF stimulation of luciferase expression from this modified reporter construct. PDGF or serum treatment both activate c-Jun N-terminal kinase (JNK), the mitogen-activated protein kinase responsible for phosphorylation and activation of c-Jun. Cotransfection of plasmids expressing dominant-negative Ras, Rac1, MEKK-1, or JNK along with the [PGS2][luciferase] reporter prevents induction by PDGF or serum, demonstrating that serum and PDGF induction of the PGS2 gene in NIH 3T3 cells requires activation of a Ras/Rac1/MEKK-1/JNK kinase/JNK signal transduction leading to phosphorylation of c-Jun. Additional cotransfection experiments with plasmids expressing dominant-negative Raf1 and ERK demonstrate that induction of PGS2 gene expression by PDGF and serum also requires activation of a Ras/Raf1/mitogen-activated protein kinase kinase (MAPKK)/ERK signal transduction pathway.
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PMID:Transcriptional regulation of prostaglandin synthase 2 gene expression by platelet-derived growth factor and serum. 894 Jan 99

Cyclic AMP-responsive element binding protein (CREB) mediates gene expression in response to cAMP stimulation. The transcriptional activity of CREB depends on both the phosphorylation of Ser133 and the recruitment of cofactor for assembly of transcriptional complex. Extensive Ser133 phosphorylation of CREB was induced during T cell activation. This phosphorylation event is essential for IL-2 gene expression. However, phosphorylation of CREB at Ser133 was not sufficient for transcriptional activity by CREB. The presence of a second signal from CD28, a potent costimulatory molecule on T cells, stimulated CREB-mediated gene expression. CD28, an effective costimulator of T cell activation and IL-2 gene expression, is shown to induce CREB activation in the presence of anti-CD3 or O-tetradecanoylphorbol 13-acetate. These two signals together stimulated a CRE-dependent reporter gene, the proliferating cell nuclear Ag promoter, and transactivation by the GAL4-CREB fusion protein. Thus optimal induction of CREB, similar to the full activation of T lymphocytes, may be mediated by two distinct signal transductions. Using the specific kinase inhibitor, one of the two pathways appeared to involve mitogen-activated protein kinase kinase but not protein kinase C, protein kinase A, or p70 S6 kinase.
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PMID:CD28-costimulation activates cyclic AMP-responsive element-binding protein in T lymphocytes. 897 78

Previous studies have shown that a mitogen activated protein (MAP) kinase (MEK)-independent signaling pathway is required by activated Raf or fibroblast-derived growth factor (FGF) for the differentiation of rat hippocampal neuronal H19-7 cells. We now demonstrate that both Raf and FGF similarly induce prolonged transcription and translation of the immediate early gene pip92 in the absence of activation of the MAP kinases (MAPKs) ERK1 and ERK2. To determine the mechanism by which this occurs and to identify novel Raf-activated signaling pathways, we investigated the induction of the pip92 promoter by both FGF and an estradiol-activated Raf-1-estrogen receptor fusion protein (deltaRaf-1:ER) in H19-7 cells. Deletion analysis of the pip92 promoter indicated that activation by the MAPK-independent pathway occurs primarily within the region containing a serum response element (SRE). Further analysis of the SRE by using a heterologous thymidine kinase promoter showed that both an Ets and CArG-like site are required. Elk1, which binds to the Ets site, was phosphorylated both in vitro and in vivo by the MAPK-independent pathway, and phosphorylation of an Elk1-GAL4 fusion protein by this pathway was sufficient for transactivation. Finally, at least two Elk1 kinases were fractionated by gel filtration, and analysis by an in-gel kinase assay revealed at least three novel Raf-activated Elk1 kinases. These results indicate that both FGF and Raf activate MAPK-independent kinases that can stimulate Elk1 phosphorylation and immediate early gene transcription.
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PMID:Raf and fibroblast growth factor phosphorylate Elk1 and activate the serum response element of the immediate early gene pip92 by mitogen-activated protein kinase-independent as well as -dependent signaling pathways. 952 98

The RBCK1 protein was recently identified as a protein kinase C-interacting protein with a new type of RBCC (RING-B-Box-Coiled-coil) region, possessing both DNA-binding and transcriptional activities unlike other proteins in the RBCC protein family (Tokunaga et al. Biochem. Biophys. Res. Commun. 244, 353-359, 1998). To identify protein motifs in the RBCC region of RBCK1 essential for the transcriptional activity, RBCK1 mutant proteins have been constructed and analyzed by using the GAL4 chimeric transcription regulator system. We have found that both of the RING-finger and the B-Box motifs are indispensable for the transcriptional activity of RBCK1. This is the first observation that these protein motifs of the RBCC protein family play a crucial role in transcriptional activation. In addition, we have examined the effect of co-expression of several protein kinases on the transcriptional activity of RBCK1. Protein kinase A (PKA) was found to enhance the activity by about eightfold, whereas both ERK (extracellular signal-regulated kinase) activator kinase 1 (MEK1) and MEK kinase 1 (MEKK1) significantly repressed the activity. Because RBCC proteins are presumed to act as a proto-oncoprotein, these results suggest that the RBCK1 protein is involved in the intracellular signaling cascades along with PKA, MEK1, and MEKK1 and mediates cell growth and differentiation.
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PMID:Transcriptional activity of RBCK1 protein (RBCC protein interacting with PKC 1): requirement of RING-finger and B-Box motifs and regulation by protein kinases. 964 38

Protein kinase C (PKC) designates a family of kinases that regulate many essential functions including cell growth and differentiation. The tight regulation of PKC activity is crucial for maintaining normal cellular proliferation and excessive activity leads to abnormal or uncontrolled cell growth. Recent reports indicate that malignant glioma cell lines express 100 to 1000-fold higher PKC activity when compared to non-neoplastic astrocytes. This high activity correlates well with the proliferation of tumor cells in vitro. We recently reported on the anti-proliferative properties of selective PKC inhibitors on the growth of U-373MG human astrocytoma cell line, and their ability to block mitogen-activated protein (MAP) kinase pathway activated by substance P (SP) neuropeptide receptor signaling via a PKC-dependent mechanism. Therefore, inhibiting PKC activity by selective PKC inhibitors may present a promising approach for improving astroglial brain tumor therapy. For this purpose, we constructed a high throughput model cell system to evaluate the efficacy of PKC inhibitors. This system is based on the measurement of light production in U-373MG cells stably transfected with the luciferase reporter gene whose expression depends on the transcriptional activation of GAL4-Elk1 fusion protein by enzyme components of the MAP kinase pathway and the upstream activation of PKC (PKC activation-->MAP kinases-->GAL4-Elk1 phosphorylation-->luciferase expression-->luciferase activity). In brief, we have demonstrated that the PKC activator 12-O-tetradecanoyl phorbol 13-acetate (TPA)-induced luciferase activity in this cell system is mediated via the MAP kinase pathway and can be blocked in the presence of MEK1 selective inhibitors (PD 098059 or U0126). We also demonstrated that TPA-induced luciferase activity in U-373MG stable clones can be blocked by PKC inhibitors (CGP 41251, Go 6976, and GF 109203X) in a concentration dependent manner. In contrast, epidermal growth factor (EGF)-induced luciferase activity, which is independent of PKC activation (Ras-->Raf-1-->MEK1-->MAP kinases-->GAL4-Elk1 phosphorylation-->luciferase expression-->luciferase activity) can only be blocked using a selective EGF receptor inhibitor (AG 1478). In conclusion, we have constructed a model cell system for the high throughput screening and identification of PKC inhibitors potentially active against astrocytoma cells in culture.
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PMID:A high throughput system for the evaluation of protein kinase C inhibitors based on Elk1 transcriptional activation in human astrocytoma cells. 991 10

Transforming growth factor-beta (TGF-beta) exerts its effects on cell proliferation, differentiation and migration in part through its modulation of extracellular matrix components, such as fibronectin and plasminogen activator inhibitor-1 (PAI-1). Although the SMAD family of proteins recently has been shown to be a key participant in TGF-beta signaling, other signaling pathways have also been shown to be activated by TGF-beta. We report here that c-Jun N-terminal kinase (JNK), a member of the MAP kinase family, is activated in response to TGF-beta in the human fibrosarcoma HT1080-derived cell line BAHgpt. Stable expression of dominant-negative forms of JNK1 and MKK4, an upstream activator of JNK, results in loss of TGF-beta-stimulated fibronectin mRNA and protein induction, while having little effect on TGF-beta-induced levels of PAI-1. The human fibronectin promoter contains three CRE elements, one of which has been shown to bind a c-Jun-ATF-2 heterodimer. Utilizing a GAL4 fusion trans-reporting system, we demonstrate a decrease in transactivating potential of GAL4-c-Jun and GAL4-ATF-2 in dominant-negative JNK1- and MKK4-expressing cells. Finally, we show that TGF-beta-induced fibronectin synthesis is independent of Smad4. These results demonstrate that TGF-beta-mediated fibronectin induction requires activation of JNK which in turn modulates the activity of c-Jun and ATF-2 in a Smad4independent manner.
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PMID:TGF-beta induces fibronectin synthesis through a c-Jun N-terminal kinase-dependent, Smad4-independent pathway. 1006

Pituitary tumor-transforming gene (PTTG) is a recently characterized oncogene that can act as a transcriptional activator. In this study, we have characterized the transactivation domain of PTTG. Transient transfection of fusion constructs containing GAL4 DNA-binding domain and different parts of PTTG indicated the transactivation domain of PTTG is located between amino acids 119 and 164. Mitogen-activated protein (MAP) kinase cascade is important in the regulation of cell growth, apoptosis, and differentiation. Therefore, we have explored the possibility that this kinase cascade plays a role in regulating PTTG transactivation function. Activation of the MAP kinase cascade by epidermal growth factor or an expression vector for a constitutively active form of the MAP kinase kinase (MEK1) led to stimulation of PTTG transactivation activity. We showed that PTTG is phosphorylated in vitro on Ser(162) by MAP kinase and that this phosphorylation site plays an essential role in PTTG transactivation function. We demonstrated that PTTG interacts directly with MEK1 through a putative SH3 domain-binding site located between amino acids 51 and 54 and that this interaction is crucial for PTTG transactivation function. In addition, we showed that activation of MAP kinase phosphorylation cascade resulted in nuclear translocation of PTTG. Together, our data establish that a growth factor-stimulated MAP kinase plays an important role in modulating PTTG function.
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PMID:Activation of mitogen-activated protein kinase cascade regulates pituitary tumor-transforming gene transactivation function. 1090 23

Intraarticular injection of dexamethasone (DEX) accelerates cartilage degradation due to the suppression of chondrocyte proliferation and extracellular matrix formation. The present study first demonstrated the interaction between DEX and TGF beta, a potent growth factor for cultured rat articular chondrocytes (CRAC), and then investigated the molecular mechanism by which DEX counteracts TGF beta-induced chondrocyte proliferation and differentiation through the regulation of AP-1 activity. DEX reduced serum-deprived and TGF beta-stimulated cell growth and [(3)H]-thymidine incorporation of CRAC. DEX also inhibited the expression of (alpha)1 type II collagen with concomitant suppression of the promoter activity. Transfection studies using a reporter vector with AP-1 responsive elements showed that DEX reduced TGF beta-activated but not basal luciferase activities. Activation of 3TP-luc, another AP-1 responsive element containing reporter was also blocked by DEX. GAL4-Elk1 studies revealed that DEX suppressed TGF beta-induced ERK activation which led to c-fos gene expression followed by increase in AP-1 complex formation, whereas the Smad pathway was not involved in DEX-dependent negative regulation of AP-1 in a reporter assay that requires FAST1-Smad2 for the activation. DEX also eliminated TGF beta-induced c-fos mRNA expression and ERK activation in Northern analysis and in vitro kinase assay, respectively. Further, DNA synthesis and transactivation of type II collagen by TGF beta were inhibited by PD98059, an inhibitor of MEK. Our results indicate that DEX suppressed TGF beta-induced chondrocyte proliferation and type II collagen expression, probably through selective inhibition of ERK integrated AP-1 activation.
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PMID:Dexamethasone inhibition of TGF beta-induced cell growth and type II collagen mRNA expression through ERK-integrated AP-1 activity in cultured rat articular chondrocytes. 1096 45

High glucose (HG) stimulates glomerular mesangial cell (MC) expression of extracellular matrix, a process involving protein kinase C (PKC) isozymes and enhanced signaling by autocrine peptides such as endothelin-1 (ET-1). The purpose of this study was to identify the specific PKC isozymes mediating the effects of HG on MC extracellular signal-regulated protein kinase (ERK1/2) signaling and alpha1(IV) collagen expression in response to ET-1. HG (30 mmol/l for 72 h) enhanced ET-1-stimulated alpha1(IV) collagen mRNA expression from 1.2 +/- 0.1-fold to 1.9 +/- 0.2-fold (P < 0.05 vs. normal glucose [NG] + ET-1), and the effect was significantly reduced by Calphostin C or the MEK (mitogen-activated protein kinase kinase) inhibitor PD98059. In transiently transfected MCs, dominant-negative (DN)-PKC-delta, -epsilon, or -zeta inhibited ET-1 activation of ERK1/2. Likewise, downstream of ERK1/2, ET-1 stimulated Elk-1-driven GAL4 luciferase activity to 11 +/- 1-fold (P < 0.002 vs. NG + ET-1) in HG, and DN-PKC-delta, -epsilon, or -zeta attenuated this response to NG levels. HG enhanced ET-1-stimulated intracellular alpha1(IV) collagen protein expression, assessed by confocal immunofluorescence imaging, showed that individual DN-PKC-delta, -epsilon, -zeta, as well as DN-PKC-alpha and -beta, attenuated the response. Thus, HG-enhanced ET-1 stimulation of alpha1(IV) collagen expression requires PKC-delta, -epsilon, and -zeta to act through an ERK1/2-dependent pathway and via PKC-alpha and -beta, which are independent of ERK1/2.
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PMID:High glucose-enhanced mesangial cell extracellular signal-regulated protein kinase activation and alpha1(IV) collagen expression in response to endothelin-1: role of specific protein kinase C isozymes. 1157 22

Transforming growth factor-beta (TGF-beta) and insulin-like growth factors (IGFs) play critical roles in the control of myogenesis. Insulin-like growth factor-binding protein-5 (IGFBP-5), by regulating the bioavailability of IGFs, is involved in controlling IGF-dependent differentiation. We investigated the effects of TGF-beta on the IGFBP-5 production induced by IGFs in mouse myoblasts. TGF-beta leads to a decrease in IGFBP-5 synthesis at both transcript and protein levels, and blocked muscle differentiation. The Smad proteins and the c-Jun N-terminal kinase (JNK) have been shown to be involved in TGF-beta signaling pathways. We provide evidence that the JNK pathway, rather than Smad proteins, is involved in the response of muscle cells to TGF-beta. This factor failed to stimulate the GAL4-Smad 2/3 transcriptional activities of the constructs used to transfect myoblasts. Moreover, stable expression of the antagonistic Smad7 did not abolish the inhibitory effect of TGF-beta on IGFBP-5 production whereas expression of a dominant-negative version of MKK4, an upstream activator of JNK, did. We also showed, using a specific inhibitor, that the p38 mitogen-activated protein kinase (p38 MAPK) was not involved in the inhibition of IGFBP-5 production. Thus, TGF-beta-mediated IGFBP-5 inhibition is independent of Smads and requires activation of the JNK signaling pathway.
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PMID:Transforming growth factor-beta inhibition of insulin-like growth factor-binding protein-5 synthesis in skeletal muscle cells involves a c-Jun N-terminal kinase-dependent pathway. 1159 9


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