Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

T-cell antigen receptor (TCR) ligation of an Lck-deficient Jurkat mutant, J.CaM1, with anti-CD3 or anti-TCR beta monoclonal antibodies failed to induce tyrosine phosphorylation and activation of p42MAPK. The same stimuli activated mitogen-activated protein (MAP) kinase in J.CaM1 cells transfected with Lck, demonstrating that Lck plays a critical role in MAP kinase activation. Utilizing immunocomplex kinase assays, we demonstrated that TCR/CD3 ligation activated a MAP kinase kinase kinase (Raf-1) as well as a MAP kinase kinase (MEK-1) in Jurkat but not in J.CaM1 cells. It was possible, however, to activate Raf-1, MEK-1, and p42MAPK in J.CaM1 cells during treatment with the phorbol ester phorbol 12-myristate 13-acetate, which activates protein kinase C (PKC). This demonstrates the presence of a PKC-dependent pathway which functions independently from Lck in MAP kinase activation. Stimulation of Jurkat cells with either anti-TCR beta or anti-CD3 monoclonal antibody failed to induce substantial tyrosine phosphorylation of Shc proteins or their association with Grb2 which forms a complex with the guanine nucleotide exchange factor hSOS. However, the same stimuli induced tyrosine phosphorylation of another putative guanine nucleotide exchange factor, p95Vav, in Jurkat but not J.CaM1 cells. Moreover, Lck was reversibly co-immunoprecipitated with p95Vav, and the stoichiometry of binding increased in anti-CD3-treated Jurkat cells. Phorbol 12-myristate 13-acetate did not induce tyrosine phosphorylation of p95Vav. These data show that the TCR activates MAP kinase by way of a signaling cascade, which depends upon Lck, and may be mediated by downstream events involving PKC or p95Vav which act on Raf-1 and MEK-1.
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PMID:The T-cell antigen receptor utilizes Lck, Raf-1, and MEK-1 for activating mitogen-activated protein kinase. Evidence for the existence of a second protein kinase C-dependent pathway in an Lck-negative Jurkat cell mutant. 751 37

Mitogen-activated protein kinases (MAPKs) are activated upon a variety of extracellular stimuli in different cells. In macrophages, colony-stimulating factor 1 (CSF-1) stimulates proliferation, while bacterial lipopolysaccharide (LPS) inhibits cell growth and causes differentiation and activation. Both CSF-1 and LPS rapidly activate the MAPK network and induce the phosphorylation of two distinct ternary complex factors (TCFs), TCF/Elk and TCF/SAP. CSF-1, but not LPS, stimulated the formation of p21ras. GTP complexes. Expression of a dominant negative ras mutant reduced, but did not abolish, CSF-1-mediated stimulation of MEK and MAPK. In contrast, activation of the MEK kinase Raf-1 was Ras independent. Treatment with the phosphatidylcholine-specific phospholipase C inhibitor D609 suppressed LPS-mediated, but not CSF-1-mediated, activation of Raf-1, MEK, and MAPK. Similarly, down-regulation or inhibition of protein kinase C blocked MEK and MAPK induction by LPS but not that by CSF-1. Phorbol 12-myristate 13-acetate pretreatment led to the sustained activation of the Raf-1 kinase but not that of MEK and MAPK. Thus, activated Raf-1 alone does not support MEK/MAPK activation in macrophages. Phosphorylation of TCF/Elk but not that of TCF/SAP was blocked by all treatments that interfered with MAPK activation, implying that TCF/SAP was targeted by a MAPK-independent pathway. Therefore, CSF-1 and LPS target the MAPK network by two alternative pathways, both of which induce Raf-1 activation. The mitogenic pathway depends on Ras activity, while the differentiation signal relies on protein kinase C and phosphatidylcholine-specific phospholipase C activation.
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PMID:Ras-dependent and -independent pathways target the mitogen-activated protein kinase network in macrophages. 779 56

12-O-Tetradecanoylphorbol 13-acetate (TPA) induces HL-60 cells to differentiate along the monocyte/macrophage pathway and stimulates expression of the extracellular adhesion protein osteopontin (OPN). In this study, the mechanism of TPA-mediated OPN mRNA expression and its relationship to differentiation were investigated. The induction of OPN mRNA by TPA was dose dependently inhibited by staurosporine (0.4-10.0 nM) and chelerythrine (0.1-5.0 microM), indicating that OPN expression requires PKC activation. Furthermore, the mitogen-activated protein kinase kinase (MAPKK) inhibitor, PD 098059 (1.0-10.0 microM), inhibited the effect of TPA in a dose-dependent fashion. Cycloheximide (10 microg/ml) ablated the induction of OPN mRNA by TPA. To determine if OPN mRNA expression was associated with a particular differentiational pathway, HL-60 cells were treated with RA, 9-cis-RA, calcitriol, or sodium butyrate. None of these agents stimulated OPN mRNA. Treatment with TPA subsequent to a 120-h pretreatment with retinoic acid (RA), 9-cis-RA, or calcitriol resulted in a potentiation of the induction of OPN mRNA. These results support a role for protein kinase C (PKC) in promoting OPN expression because each of these agents increased PKC levels. An hOPN promoter/reporter construct was responsive to TPA, indicating that this effect is at the level of transcription. Thus, TPA-stimulated transcription of the OPN gene apparently occurs via a PKC/MAPK-dependent mechanism that is independent of that associated with differentiation and is not dependent on the maturational state of these cells.
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PMID:Stimulation of osteopontin mRNA expression in HL-60 cells is independent of differentiation. 922 25

The GT1-1 GnRH neuronal cell lines exhibit highly differentiated properties of GnRH neurons. We have used GT1-1 cells to study the roles of norepinephrine (NE), membrane depolarization, calcium influx, and phorbol esters in the regulation of mitogen-activated protein (MAP) kinase. NE, which is known to stimulate the release of GnRH, induced MAP kinase activity, the tyrosine phosphorylation of MAP kinase, and MAP kinase kinase activity. Forskolin led to activation of MAP kinase comparable with that induced by NE, and a selective inhibitor of cAMP-dependent protein kinase, H8, attenuated the NE-induced activation of MAP kinase. On the other hand, elimination of extracellular calcium by EGTA completely blocked NE-induced tyrosine phosphorylation of MAP kinase, and a selective inhibitor of calcium/calmodulin-dependent protein kinase, KN-62, attenuated the NE-induced activation of MAP kinase. Furthermore, depolarization of GT1-1 cells with 75 mM KCl, 10 microM BayK 8644, or 1 microM calcium ionophore (A23187) induced rapid tyrosine phosphorylation of MAP kinase. The omission of calcium from the extracellular medium completely abolished these effects of tyrosine phosphorylation of MAP kinase. Phorbol 12-myristate 13-acetate (PMA) also induced MAP kinase activity, but pretreatment of the cultured cells with PMA to down-regulate protein kinase C did not abolish the activation of MAP kinase by NE. In addition, although phosphorylation of Raf-1 kinase was stimulated by PMA, this phosphorylation was not induced by either NE or A23187. These results demonstrate that NE activates MAP kinase directly in GT1-1 cells, and that the effect of NE is mediated by increase in the cAMP level and by calcium influx, but not by PMA-sensitive protein kinase C or Raf-1 kinase.
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PMID:Norepinephrine stimulates mitogen-activated protein kinase activity in GT1-1 gonadotropin-releasing hormone neuronal cell lines. 938 11

Erythropoietin (EPO), a major regulator of erythroid progenitor cells, is essential for the survival, proliferation, and differentiation of immature erythroid cells. To gain insight into the molecular mechanism by which EPO functions, we analyzed the activation of Jun N-terminal kinases (JNKs) and extracellular signal-regulated kinases (ERKs) in HCD-57 cells, a murine erythroid progenitor cell line that requires EPO for survival and proliferation. Withdrawal of EPO from the cell culture medium resulted in sustained activation of JNKs plus p38 MAP kinase, and inactivation of ERKs, preceding apoptosis of the cells. Addition of EPO to the EPO-deprived cells caused activation of ERKs accompanied by inactivation of JNKs and p38 MAP kinase and rescued the cells from apoptosis. Phorbol 12-myristate 13-acetate, which activated ERKs by a different mechanism, also suppressed the activation of JNKs and significantly retarded apoptosis of the cells caused by withdrawal of EPO. Furthermore, MEK inhibitor PD98059, which inhibited activation of ERKs, caused activation of JNKs, whereas suppression of JNK expression by antisense oligonucleotides and inhibition of p38 MAP kinase by SB203580 caused attenuation of the apoptosis that occurs upon withdrawal of EPO. Finally, the activation of JNKs and p38 MAP kinase and concurrent inactivation of ERKs upon withdrawal of EPO were also observed in primary human erythroid colony-forming cells. Taken together, the data suggest that activation of ERKs promotes cell survival, whereas activation of JNKs and p38 MAP kinase leads to apoptosis and EPO functions by controlling the dynamic balance between ERKs and JNKs.
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PMID:Distinct roles of JNKs/p38 MAP kinase and ERKs in apoptosis and survival of HCD-57 cells induced by withdrawal or addition of erythropoietin. 1187 10

Phorbol 12-myristate 13-acetate (PMA)-induced differentiation of human erythroleukemic K562 cells is characterized by growth arrest, morphological change, and expression of megakaryocyte-specific proteins. We examined the possible involvement of cell cycle regulators with PMA-induced growth arrest and megakaryocytic differentiation of K562 cells. The concentrations of cyclin D1 and p21Waf1/Cip1 were dramatically increased, whereas those of cyclin B1 and cdc2 were decreased, by PMA treatment. The concentrations of most cyclin-dependent kinases (Cdk2, Cdk4, and Cdk6), however, were unchanged by PMA treatment. PD98059, a specific inhibitor of MEK1, partially prevented the increase in cyclin D1 caused by PMA and fully reversed the down-regulation of cyclin B1 protein seen in response to PMA treatment. Thus, it is demonstrated here that the PMA-mediated changes of cyclin D1 and B1 are the result of a persistent increase in extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) activity.
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PMID:ERK/MAPK pathway is required for changes of cyclin D1 and B1 during phorbol 12-myristate 13-acetate-induced differentiation of K562 cells. 1068 62

Phosphodiesterase 4D5 is the sole PDE4D cAMP phosphodiesterase isoform expressed in human aortic smooth muscle cells (HASMC). Phorbol 12-myristate 13-acetate (PMA) challenge of HASMC rapidly activated PDE4D5 through a process ablated by the mitogen-activated protein kinase kinase inhibitor PD98059. PMA elicited an inhibitory effect on PDE4D5 activity in HASMC treated with the cyclooxygenase (COX) inhibitor indomethacin, the COX-2 selective inhibitor NS-398, the phospholipase A(2) inhibitor quinacrine, and the cAMP-dependent protein kinase A (PKA) inhibitor H89. PMA challenge of COS-1 cells elicited the rapid inhibition and phosphorylation of both recombinant and endogenous PDE4D5 in a manner ablated by PD98059 and not seen in S651A mutant PDE4D5. PMA promoted the generation of PGE(2) in the medium of HASMC and caused activation of both extracellular signal-regulated kinase (ERK) and PKA through a process ablated by indomethacin, NS-398, quinacrine, and PD98059. Exogenous prostaglandin (PG) E(2) increased cAMP levels and activated PKA in HASMC. COX-2 was expressed in HASMC but not in COS-1 cells. Forskolin challenge of COS-1 cells activated PDE4D5 by causing the PKA-mediated phosphorylation of Ser126 as detected using a novel phosphospecific antiserum. PMA challenge of HASMC elicited phosphorylation of the stimulatory PKA-specific phosphorylation site, Ser126 in PDE4D5 in a manner ablated by PD98059, indomethacin, and H89. We propose that, in HASMC, PMA activates PDE4D5 through an ERK-controlled autocrine mechanism. This involves PGE(2) generation, which causes activation of adenylyl cyclase, allowing PKA to elicit net activation of PDE4D5 by phosphorylation at Ser126.
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PMID:Phorbol 12-myristate 13-acetate triggers the protein kinase A-mediated phosphorylation and activation of the PDE4D5 cAMP phosphodiesterase in human aortic smooth muscle cells through a route involving extracellular signal regulated kinase (ERK). 1164 39

To better understand the intracellular signaling mechanism that causes the association of insulin resistance and hyperlipidemia with cardiovascular diseases, we specifically looked at the ability of lysophosphatidylcholine (lysoPC) to inhibit the Akt activation induced by insulin in cultured rat aortic vascular smooth muscle cells. LysoPC inhibited the insulin-induced phosphorylation of Akt at Ser473, and the inhibition was concentration dependent. Phorbol 12-myristate 13-acetate (PMA), a protein kinase C (PKC) activator, inhibited the insulin-induced phosphorylation of Akt. LysoPC stimulated PKC phosphorylation at Ser660, which was inhibited by the PKC inhibitor GF109203X. The PKC-alpha/beta-selective inhibitor Go6976 also blocked the PMA- and lysoPC-induced inhibition of Akt phosphorylation by insulin. PKC-alpha, but not PKC-beta, is expressed in vascular smooth muscle cells, and overexpression of PKC-alpha, but not PKC-beta or PKC-delta, inhibited insulin-induced Akt activation. LysoPC rapidly stimulated PKC-alpha translocation to the membrane. In contrast, pretreatment with the p42/44 mitogen-activated protein kinase kinase inhibitor PD98059 or the p38 mitogen-activated protein kinase inhibitor SB203580 did not block the lysoPC-induced inhibition of Akt phosphorylation by insulin. In addition, lysoPC inhibited the insulin-induced tyrosine phosphorylation of insulin receptor substrate (IRS)-1 but not that of the insulin receptor beta subunit or insulin binding. PMA treatment or PKC-alpha overexpression also inhibited the tyrosine phosphorylation of IRS-1. From these data, we conclude that lysoPC negatively regulates the insulin signal at the point of IRS-1 through PKC-alpha in the vasculature, which may explain the association of hyperlipidemia with hyperinsulinemia in cardiovascular diseases.
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PMID:Lysophosphatidylcholine inhibits insulin-induced Akt activation through protein kinase C-alpha in vascular smooth muscle cells. 1188 99

We have recently shown that the Ras-Raf-MEK-ERK and phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathways can cross-talk in the human breast cancer cell line MCF-7. High Raf activity induces growth arrest and differentiation in these cells, whereas high PI3K/Akt activity correlates with cell survival and proliferation. Here we show that the Raf-Akt cross-talk is regulated in a concentration- and ligand-dependent manner. High doses of insulin-like growth factor I (IGF-I) activate Akt quickly and strongly enough to suppress Raf kinase activity via phosphorylation of Ser-259, whereas low doses of IGF-I do not trigger this cross-talk but are still mitogenic. Phorbol 12-myristate 13-acetate, a differentiation-inducing stimulus, potently activates the Ras-Raf-MEK-ERK pathway but only weakly activates PI3K/Akt and does not trigger the cross-talk. Thus, the herein analyzed parameters such as ligand type, concentration, and time course may contribute to the cellular response of either proliferation or differentiation. This is highly relevant to understanding cellular transformation and may be of use in areas like tissue engineering.
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PMID:Regulation of Raf-Akt Cross-talk. 1204 82

Angiotensin-converting enzyme (ACE) is an enzyme that plays a major role in vasoactive peptide metabolism, and it has been implicated in various cardiovascular diseases. Phorbol 12-myristate 13-acetate (PMA), a protein kinase C activator, has been shown to increase ACE mRNA at the transcriptional level in human umbilical vein endothelial cells. We have investigated the transcriptional mechanism involved in protein kinase C induction of the ACE gene. Deletion and transfection analyses have revealed that two regions are required for PMA-inducible gene expression. The first is a G+C-rich region located in the proximal ACE promoter bearing overlapping consensus recognition sequences for stimulatory protein-1 (Sp1) and early growth response gene 1 (Egr-1). Electrophoretic mobility shift assay and supershift experiments have shown that Egr-1 is present in the specific nucleoprotein complex induced by PMA in human umbilical vein endothelial cells. The second region is located in the distal ACE promoter. DNase I footprinting analysis restricted this region to a 21-bp element containing a cAMP-responsive element/12-O-tetradecanoylphorbol 13-acetate-responsive element sequence. Electrophoretic mobility shift assays and supershift analyses have revealed that activating protein 1 (AP-1) is the transcription factor binding the cAMP-responsive element/12-O-tetradecanoylphorbol 13-acetate-responsive element located in the ACE promoter after PMA stimulation. Mutations of either Egr-1 or AP-1 binding sites partially abrogate ACE expression induced by PMA, whereas mutation of both sites totally abrogates PMA-induced ACE expression. Treatment of cells with PD98059, a mitogen-activated protein kinase kinase-1-specific inhibitor, inhibited PMA-induced ACE expression. Our results demonstrate that the two transcription factors, Egr-1 and AP-1, are involved in the PMA-induced ACE transcriptional activation in human endothelial cells via the activation of the extracellular signal-regulated kinase 1/2 signaling pathway.
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PMID:Phorbol ester induction of angiotensin-converting enzyme transcription is mediated by Egr-1 and AP-1 in human endothelial cells via ERK1/2 pathway. 1243 34


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