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)

Monocyte chemoattractant protein-1 (MCP-1), a member of the C-C subfamily of chemokines, is important for the local recruitment of leukocytes to sites of inflammatory challenge. Here, we investigated endothelial signaling pathways involving members of the mitogen-activated protein (MAP) kinase superfamily and studied their role for MCP-1 expression in endothelium. We show that tumor necrosis factor-alpha (TNF-alpha), a potent inflammatory activator of endothelium, leads to activation of MAP kinases ERK, p38, and JNK in human umbilical vein endothelial cells (HUVEC). Contribution of MAP kinase pathways to TNF-alpha-induced synthesis of endothelial MCP-1 was then studied by pharmacologic inhibition and transient expression of dominant negative or constitutively active kinase mutants using flow cytometry, Northern blot, and luciferase reporter gene assays. Inhibition of Raf/MEK/ERK or SEK/JNK pathways had no significant effect on MCP-1 levels, whereas blocking the MKK6/p38 pathway by p38 inhibitors SB203580 or SB202190 or by a dominant negative mutant of MKK6, the upstream activator of p38, strongly inhibited TNF-alpha-induced expression of MCP-1. Consistent with that finding, expression of wild-type or constitutively active MKK6 significantly enhanced the effect of limiting TNF-alpha concentrations on MCP-1 synthesis. These data suggest a crucial role for the MKK6/p38 stress kinase cascade in TNF-alpha-mediated endothelial MCP-1 expression.
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PMID:The MKK6/p38 stress kinase cascade is critical for tumor necrosis factor-alpha-induced expression of monocyte-chemoattractant protein-1 in endothelial cells. 992 Aug 34

Because both the brain natriuretic peptide (BNP) gene and the cytokine interleukin-1beta (IL-1beta) are induced in the infarcted myocardium, localized production of IL-1beta may regulate the BNP gene. We tested whether (1) IL-1beta regulates the human BNP promoter, (2) cis elements in the proximal promoter respond to IL-1beta, and (3) mitogen-activated protein kinase (MAPK) signaling pathways [p42/44, c-jun (JNK) and p38 kinase] are involved. We transferred the hBNP promoter coupled to a luciferase reporter gene or constructs with mutations in the proximal promoter GATA and M-CAT elements into neonatal rat ventricular myocytes and treated the cells with IL-1beta for 24 hours. IL-1beta-stimulated hBNP luciferase activity was eliminated by pretreatment with the transcription inhibitor actinomycin D. Both the p38 kinase inhibitor SB205380 (SB) and cotransfection of a dominant-negative mutant of p38 kinase reduced IL-1beta stimulation of the hBNP promoter. Dominant-negative mutants of Ras and Rac inhibited IL-1beta-stimulated hBNP luciferase activity by 64% and 90%, respectively. Constitutively active forms of Rac and MKK6, the immediate upstream activator of p38, were stimulatory; however, only the effect of MKK6 was inhibited by SB. Neither the p42/44 nor the JNK pathway was involved in the action of IL-1beta. Both IL-1beta and MKK6 activation of the hBNP promoter were partially reduced when the promoter contained a mutated M-CAT element. In summary, (1) IL-1beta is a transcriptional activator of the hBNP promoter; (2) IL-1beta acts through a Ras-dependent pathway not coupled to activation of p42/44 MAPK or JNK; (3) IL-1beta acts through a Rac-dependent pathway, but the downstream effector is not known; and (4) IL-1beta activation of p38 kinase is partially involved in regulation of the hBNP promoter, targeting the proximal M-CAT element.
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PMID:Interleukin-1beta regulation of the human brain natriuretic peptide promoter involves Ras-, Rac-, and p38 kinase-dependent pathways in cardiac myocytes. 993 Nov 18

Several extracellular stimuli mediated by G protein-coupled receptors activate c-fos promoter. Recently, we and other groups have demonstrated that signals from G protein-coupled receptors stimulate mitogen-activated protein kinases (MAPKs), extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 MAPK. The activation of these three MAPKs is mediated in part by the G protein betagamma subunit (Gbetagamma). In this study, we characterized the signals from Gbetagamma to c-fos promoter using transient transfection of c-fos luciferase into human embryonal kidney 293 cells. Activation of m2 muscarinic acetylcholine receptor and overexpression of Gbetagamma, but not constitutively active Galphai2, stimulated c-fos promoter activity. The c-fos promoter activation by m2 receptor and Gbetagamma was inhibited by beta-adrenergic receptor kinase C-terminal peptide (betaARKct), which functions as a Gbetagamma antagonist. MEK1 inhibitor PD98059 and kinase-deficient mutant of JNK kinase, but not p38 MAPK inhibitor SB203580, attenuated the m2 receptor- and Gbetagamma-induced c-fos promoter activation. Activated mutants of Ras and Rho stimulated the c-fos promoter activity, and the dominant negative mutants of Ras and Rho inhibited the c-fos promoter activation by m2 receptor and Gbetagamma. Moreover, c-fos promoter activation by m2 receptor, Gbetagamma, and active Rho, but not active Ras, was inhibited by botulinum C3 toxin. These data indicated that both Ras- and Rho-dependent signaling pathways are essential for c-fos promoter activation mediated by Gbetagamma.
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PMID:Activation of c-fos promoter by Gbetagamma-mediated signaling: involvement of Rho and c-Jun N-terminal kinase. 1005 39

In this study we examined the contribution of MAPK1 and 2 [also known as extracellular signal-regulated kinases (ERK)-1 and 2] to the induction of zif268 mRNA in PC12D cells by using two methods to block the activation of these kinases. In one set of experiments, we inhibited the activation of MAPK by pretreating cells with PD098059, a specific inhibitor of MEK (MAPKK), the immediate upstream activator of MAPK. In the second set of experiments, we blocked the activation of MAPK by overexpressing N17Ras, a dominant-negative form of Ha-Ras. These two approaches yielded similar results and showed that inhibition of MAPK blocks less than half of the induction of zif268 mRNA by NGF. Much of the residual induction of zif268 mRNA is blocked by low concentrations of wortmannin, an inhibitor of phosphatidylinositol (PI) 3-kinase. Since PI 3-kinase was previously shown to function upstream in epidermal growth factor (EGF)-mediated activation of c-Jun N-terminal kinase (JNK), and JNK is known to phosphorylate and activate transcription factors that regulate the expression of zif268, we investigated the role of JNK in the induction of zif268 mRNA by NGF. Stimulation of PC12D cells with NGF weakly activates JNK, but this activation is enhanced rather than inhibited by pretreatment with wortmannin, suggesting that JNK does not function downstream of PI 3-kinase in the induction of zif268 mRNA. A role for JNK in the induction of the zif268 gene is indicated, however, by the fact that cotransfection of expression vectors encoding JIP-1 or the JNK binding domain of JIP-1, which act as dominant-negative inhibitors of JNK, partially blocks the NGF-mediated induction of a luciferase reporter gene linked to the zif268 promoter. Together, these results suggest that MAPK, PI-3 kinase and JNK each play a role in the induction of zif268 gene expression by NGF in PC12D cells.
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PMID:Nerve growth factor induces zif268 gene expression via MAPK-dependent and -independent pathways in PC12D cells. 1005 43

Myocardial hypertrophy is associated with increased basal glucose metabolism. Basal glucose transport into cardiac myocytes is mediated by the GLUT1 isoform of glucose transporters, whereas the GLUT4 isoform is responsible for regulatable glucose transport. Treatment of neonatal cardiac myocytes with the hypertrophic agonist 12-O-tetradecanoylphorbol-13-acetate or phenylephrine increased expression of Glut1 mRNA relative to Glut4 mRNA. To study the transcriptional regulation of GLUT1 expression, myocytes were transfected with luciferase reporter constructs under the control of the Glut1 promoter. Stimulation of the cells with 12-O-tetradecanoylphorbol-13-acetate or phenylephrine induced transcription from the Glut1 promoter, which was inhibited by cotransfection with the mitogen-activated protein kinase phosphatases CL100 and MKP-3. Cotransfection of the myocytes with constitutively active versions of Ras and MEK1 or an estrogen-inducible version of Raf1 also stimulated transcription from the Glut1 promoter. Hypertrophic induction of the Glut1 promoter was also partially sensitive to inhibition of the phosphatidylinositol 3-kinase pathway and was strongly inhibited by cotransfection with dominant-negative Ras. Thus, Ras activation and pathways downstream of Ras mediate induction of the Glut1 promoter during myocardial hypertrophy.
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PMID:Transcriptional activation of the glucose transporter GLUT1 in ventricular cardiac myocytes by hypertrophic agonists. 1008 48

Homologous regulation of GnRH receptor (GnRHR) gene expression is an established mechanism for controlling the sensitivity of gonadotropes to GnRH. We have found that expression of the GnRHR gene in the gonadotrope-derived alpha T3-1 cell line is mediated by a tripartite enhancer that includes a consensus activator protein-1 (AP-1) element, a binding site for SF-1 (steroidogenic factor-1), and an element we have termed GRAS (GnRHR-activating sequence). Further, in transgenic mice, approximately 1900 b.p. of the murine GnRHR gene promoter are sufficient for tissue-specific expression and GnRH responsiveness. The present studies were designed to further delineate the molecular mechanisms underlying GnRH regulation of GnRHR gene expression. Vectors containing 600 bp of the murine GnRHR gene promoter linked to luciferase (LUC) were transiently transfected into alpha T3-1 cells and exposed to treatments for 4 or 6 h. A GnRH-induced, dose-dependent increase in LUC expression of the -600 promoter was observed with maximal induction of LUC noted at 100 nM GnRH. We next tested the ability of GnRH to stimulate expression of vectors containing mutations in each of the components of the tripartite enhancer. GnRH responsiveness was lost in vectors containing mutations in AP-1. Gel mobility shift data revealed binding of fos/jun family members to the AP-1 element of the murine GnRHR promoter. Treatment with GnRH or phorbol-12-myristate-13-acetate (PMA) (100 nM), but not forskolin (10 microM), increased LUC expression, which was blocked by the protein kinase C (PKC) inhibitor, GF109203X (100 nM), and PKC down-regulation (10 nM PMA for 20 h). In addition, a specific MEK1/MEK2 inhibitor, PD98059 (60 microM), reduced the GnRH and PMA responses whereas the L-type voltage-gated calcium channel agonist, +/- BayK 8644 (5 microM), and antagonist, nimodipine (250 nM), had no effect on GnRH responsiveness. Furthermore, treatment of alpha T3-1 cells with 100 nM GnRH stimulated phosphorylation of both p42 and p44 forms of extracellular signal-regulated kinase (ERK), which was completely blocked with 60 microM PD98059. We suggest that GnRH regulation of the GnRHR gene is partially mediated by an ERK-dependent activation of a canonical AP-1 site located in the proximal promoter of the GnRHR gene.
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PMID:Homologous regulation of the gonadotropin-releasing hormone receptor gene is partially mediated by protein kinase C activation of an activator protein-1 element. 1019 63

We addressed the mechanisms of restoration of cell surface proteinase-activated receptor-1 (PAR-1) by investigating thrombin-activated signaling pathways involved in PAR-1 re-expression in endothelial cells. Exposure of endothelial cells transfected with PAR-1 promoter-luciferase reporter construct to either thrombin or PAR-1 activating peptide increased the steady-state PAR-1 mRNA and reporter activity, respectively. Pretreatment of reporter-transfected endothelial cells with pertussis toxin or co-expression of a minigene encoding 11-amino acid sequence of COOH-terminal Galphai prevented the thrombin-induced increase in reporter activity. Pertussis toxin treatment also prevented thrombin-induced MAPK phosphorylation, indicating a role of Galphai in activating the downstream MAPK pathway. Expression of constitutively active Galphai2 mutant or Gbeta1gamma2 subunits increased reporter activity 3-4-fold in the absence of thrombin stimulation. Co-expression of dominant negative mutants of either Ras or MEK1 with the reporter construct inhibited the thrombin-induced PAR-1 expression, whereas constitutively active forms of either Ras or MEK1 activated PAR-1 expression in the absence of thrombin stimulation. Expression of dominant negative Src kinase or inhibitors of phosphoinositide 3-kinase also prevented the MAPK activation and PAR-1 expression. We conclude that thrombin-induced activation of PAR-1 mediates PAR-1 expression by signaling through Gi1/2 coupled to Src and phosphoinositide 3-kinase, and thereby activating the downstream Ras/MAPK cascade.
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PMID:Thrombin induces proteinase-activated receptor-1 gene expression in endothelial cells via activation of Gi-linked Ras/mitogen-activated protein kinase pathway. 1022 46

Using primary fibroblasts in culture, we have investigated the signal transduction mechanisms by which phorbol esters, a class of tumor promoters, activate the 9E3 gene and its chemokine product the chicken chemotactic and angiogenic factor. This gene is highly stimulated by phorbol 12,13-dibutyrate (PDBu) via three pathways: (i) a small contribution through protein kinase C (the commonly recognized pathway for these tumor promoters), (ii) a contribution involving tyrosine kinases, and (iii) a larger contribution via pathways that can be interrupted by dexamethasone. All three of these pathways converge into the mitogen-activated protein kinases, MEK1/ERK2. Using a luciferase reporter system, we show that although both the AP-1 and PDRIIkB (a NFkappaB-like factor in chickens) response elements are capable of activation in these normal cells, regions of the 9E3 promoter containing them are unresponsive to PDBu stimulation. In contrast, we show for the first time that activation by PDBu occurs through a segment of the promoter containing Elk1 response elements; deletion and mutation of these elements abrogates 9E3/chicken chemotactic and angiogenic factor expression. Electrophoretic mobility shift assays and functional studies using PathDetect systems show that stimulation of the cells by phorbol esters leads to activation of the Elk1 transcription factor, which binds to its element in the 9E3 promoter.
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PMID:Activation of the 9E3/cCAF chemokine by phorbol esters occurs via multiple signal transduction pathways that converge to MEK1/ERK2 and activate the Elk1 transcription factor. 1033 36

Gastrin (G17) has a CCKB receptor-mediated growth-promoting effect on the AR42J rat acinar cell line that is linked to induction of both mitogen-activated protein kinase (MAPK) and c-fos gene expression. We investigated the mechanisms that regulate the growth factor action of G17 on the rat pituitary adenoma cell line GH3. Both AR42J and GH3 cells displayed equal levels of CCKB receptor expression and similar binding kinetics of 125I-labeled G17. G17 stimulation of cell proliferation was identical in both cell lines. G17 stimulation of GH3 cell proliferation was completely blocked by the CCKB receptor antagonist D2 but not by the MEK inhibitor PD-98059 or the protein kinase C inhibitor GF-109203X, which completely inhibited G17 induction of AR42J cell proliferation. G17 induced a c-fos SRE-luciferase reporter gene plasmid more than fourfold in the AR42J cells, whereas it had no effect in the GH3 cells. In contrast to what we observed in the AR42J cells, G17 failed to stimulate MAPK activation and Shc tyrosyl phosphorylation and association with the adapter protein Grb2. Epidermal growth factor induced the MAPK pathway in the GH3 cells, demonstrating the integrity of this signaling system. G17 induced Ca2+ mobilization in both the GH3 and AR42J cells. The calmodulin inhibitor N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide inhibited AR42J cell proliferation by 20%, whereas it completely blocked G17 induction of GH3 cell growth. The Ca2+ ionophore ionomycin stimulated GH3 cell proliferation to a level similar to that observed in response to G17, but it had no effect on AR42J cell proliferation. Thus there are cell type specific differences in the requirement of the MAPK pathway for the growth factor action of G17. Whereas in the AR42J cells G17 stimulates cell growth through activation of MAPK and c-fos gene expression, in the GH3 cells, G17 fails to activate MAPK, and it induces cell proliferation through Ca2+-dependent signaling pathways. Furthermore, induction of Ca2+ mobilization in the AR42J cells appears not to be sufficient to sustain cell proliferation.
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PMID:Cell type-specific requirement of the MAPK pathway for the growth factor action of gastrin. 1036 39

Angiotensin (Ang) II stimulates proliferation of vascular smooth muscle cells (VSMC) via its specific receptor AT1 subtype, possibly leading to atherosclerosis in hypertension. On the other hand, a cytokine interferon (IFN)-gamma has been shown to have an anti-atherosclerotic effect. In the present study, we examined a possible role of IFN-gamma in AT1 receptor gene regulation in VSMC. A firefly luciferase expression vector driven by the rat AT1a receptor gene promoter ( approximately 3.2 kb) was transfected into the cultured rat VSMC, and luciferase expression was determined to estimate the transcription function of the AT1a receptor gene promoter. RT-PCR was also carried out to determine mRNA expression of AT1a receptor in VSMC. IFN-gamma treatment decreased AT1a receptor mRNA expression as well as luciferase expression in a dose-dependent manner. The analysis with deletion DNA fragments showed that the IFN-responsive element was located between -987 and -331 positions, where multiple GAS (gamma interferon activated site)-like elements were identified. The expression suppression was reversed by either a MAPKK inhibitor PD98059 or a Jak-2 inhibitor AG-490. These results suggest that IFN-gamma can inhibit AT1 receptor expression at gene transcription level, and that the transcription suppression is dependent on MAP kinase and Jak-2. Inhibition of AT1a receptor expression may possibly be implicated in the anti-atherosclerotic action of IFN-gamma in VSMC.
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PMID:Transcriptional suppression of rat angiotensin AT1a receptor gene expression by interferon-gamma in vascular smooth muscle cells. 1046 2


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