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)

To evaluate a possible mechanism for the chronic regulation of MAPK/ERK kinase-1 (MEK-1) and p42 mitogen-activated protein kinase (MAPK) we studied the long-term effects of the G-protein-coupled receptor agonist endothelin-1 (ET-1) and the protein tyrosine kinase-coupled receptor agonist platelet-derived growth factor BB (PDGF BB) on MEK-1 and p42 MAPK in glomerular mesangial cells (GMCs). ET-1 and PDGF BB led to a time-dependent increase in MEK-1 mRNA expression without altering p42 MAPK mRNA levels. The effect of ET-1 and PDGF BB on MEK-1 mRNA expression was maximal after 24 h (3.3-fold) or 6 h (2.9-fold). Furthermore, the effect of ET-1 and PDGF BB on MEK-1 mRNA expression was additive (4.2-fold after 6 h) and was inhibited by actinomycin D (5 micrograms/ml). Cycloheximide (10 micrograms/ml) inhibited MEK-1 mRNA induction but stimulated p42 MAPK mRNA expression in both the absence and the presence of ET-1 and/or PDGF BB. The ET-1 and PDGF BB-induced increase in MEK-1 mRNA was accompanied by sustained enhancement of both p45 MEK protein expression after 12 h and by elevation of p42 MAPK activity for up to 24 h. We conclude that, in GMCs, MEK-1 acts like a delayed-early gene, whereas p42 MAPK resembles an immediate-early gene. MEK-1 mRNA and protein levels, as well as p42 MAPK activity, can be chronically regulated by both a seven-transmembrane domain receptor-coupled peptide such as ET-1 and by an agonist binding to a receptor with intrinsic protein tyrosine kinase activity, such as PDGF BB.
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PMID:ET-1 and PDGF BB induce MEK mRNA and protein expression in mesangial cells. 858 80

Stimulation of Rat-1 cells with lysophosphatidic acid (LPA) or epidermal growth factor (EGF) results in a biphasic, sustained activation of extracellular signal-regulated kinase 1 (ERK1). Pretreatment of Rat-1 cells with either cycloheximide or sodium orthovanadate had little effect on the early peak of ERK1 activity but potentiated the sustained phase. Cycloheximide also potentiated ERK1 activation in Rat-1 cells expressing DeltaRaf-1:ER, an estradiol-regulated form of the oncogenic, human Raf-1. Since cycloheximide did not potentiate MEK activity but abrogated the expression of mitogen-activated protein kinase phosphatase (MKP-1) normally seen in response to EGF and LPA, we speculated that the level of MKP-1 expression may be an important regulator of ERK1 activity in Rat-1 cells. Inhibition of LPA-stimulated MEK and ERK activation with PD98059 and pertussis toxin, a selective inhibitor of Gi-protein-coupled signaling pathways, reduced LPA-stimulated MKP-1 expression by only 50%, suggesting the presence of additional MEK- and ERK-independent pathways for MKP-1 expression. Specific activation of the MEK/ERK pathway by DeltaRaf-1:ER had little or no effect on MKP-1 expression, suggesting that activation of the Raf/MEK/ERK pathway is necessary but not sufficient for MKP-1 expression in Rat-1 cells. Activation of PKC played little part in growth factor-stimulated MKP-1 expression, but LPA- and EGF-induced MKP-1 expression was blocked by buffering [Ca2+]i, leading to a potentiation of the sustained phase of ERK1 activation without potentiating MEK activity. In Rat-1DeltaRaf-1:ER cells, we observed a strong synergy of MKP-1 expression when cells were stimulated with estradiol in the presence of ionomycin, phorbol 12-myristate 13-acetate, or okadaic acid under conditions where these agents did not synergize for ERK activation. These results suggest that activation of the Raf/MEK/ERK pathway is insufficient to induce expression of MKP-1 but instead requires other signals, such as Ca2+, to fully reconstitute the response seen with growth factors. In this way, ERK-dependent and -independent signals may regulate MKP-1 expression, the magnitude of sustained ERK1 activity, and therefore gene expression.
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PMID:Regulation of mitogen-activated protein kinase phosphatase-1 expression by extracellular signal-related kinase-dependent and Ca2+-dependent signal pathways in Rat-1 cells. 914 52

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

It has been well established that hepatocyte growth factor (HGF) induces branching tubule formation of Madin-Darby canine kidney (MDCK) cells cultured in collagen gel. Tubulogenesis per se requires the involvement of cell proliferation, migration, focalization proteolysis, cell-cell interaction and differentiation. However, signaling pathways and proteins involved in HGF-induced tubulogenesis by MDCK cells have not been thoroughly studied. Because cell-matrix interactions play important roles in tubulogenesis, we analyzed whether HGF altered the expression of extracellular matrix receptor (alpha2, alpha3, beta1 and alphavbeta3 integrin). We found that among those proteins examined, alpha2beta1 integrin levels were enhanced by HGF. HGF-induced upregulation of alpha2beta1 integrin was mediated via upregulation of alpha2 integrin mRNA abundance. Cycloheximide blocked the HGF-induced increase in alpha2 integrin mRNA expression. To understand the signaling pathways leading to an HGF-induced increase in alpha2beta1 integrin levels, PD98059 (MEK1 inhibitor), LY294002 (PI3-kinase inhibitor), and GF109203X (PKC inhibitor) were used. We found that PD98059 blocked the HGF-induced increase in alpha2beta1 integrin expression. Furthermore, 5E8 (specific anti-alpha2beta1 integrin antibody) was employed to elucidate the potential role of HGF-induced upregulation of alpha2beta1 integrin in branching morphogenesis. 5E8 did not alter HGF-induced scattering effects but disrupted HGF-induced branching tubulogenesis in collagen gel via inhibition of cell-cell interactions and growth. Taken together, HGF upregulates alpha2beta1 integrin expression via an indirect pathway, the results of which contribute to the regulation of cell-cell interactions and cell growth during branching morphogenesis in collagen gel.
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PMID:Hepatocyte growth factor upregulates alpha2beta1 integrin in Madin-Darby canine kidney cells: implications in tubulogenesis. 1206 1

Aldosterone and glucocorticoids (GCs) stimulate Na(+) reabsorption in the collecting ducts by increasing the activity of the epithelial Na(+) channel (ENaC). Our laboratory has used Madin-Darby canine kidney-C7 cells to demonstrate that this effect is associated with an increase in alpha-ENaC gene transcription (Mick VE, Itani OA, Loftus RW, Husted RF, Schmidt TJ, and Thomas CP, Mol Endocrinol 15: 575-588, 2001). Cycloheximide (CHX) superinduced the GC-stimulated alpha-ENaC expression in a dose-dependent manner, but had no effect on basal or aldosterone-stimulated alpha-ENaC expression, whereas anisomycin inhibited basal and corticosteroid-stimulated alpha-ENaC expression. The superinduction of alpha-ENaC expression was also seen with hypotonicity, was blocked by RU-38486, and was independent of protein synthesis. CHX had no effect on alpha-ENaC mRNA half-life, confirming that its effect was via an increase in alpha-ENaC transcription. The effect of CHX and hypotonicity on alpha-ENaC expression was abolished by SB-202190, indicating an effect mediated via p38 MAPK. Consistent with this scheme, CHX increased pp38 and MKK6, an upstream activator of p38, stimulated alpha-ENaC promoter activity. These data confirm a model in which CHX activates p38 in Madin-Darby canine kidney-C7 cells to increase alpha-ENaC gene transcription in a GC-dependent manner.
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PMID:Cycloheximide increases glucocorticoid-stimulated alpha -ENaC mRNA in collecting duct cells by p38 MAPK-dependent pathway. 1250 61

Basic fibroblast growth factor (bFGF, FGF-2) is one of the potent mitogens for periodontal ligament (PDL) cells. However, the role of bFGF on the matrix metalloproteinase-3 (MMP-3) expression in PDL cells is unknown. In this study, the effect of bFGF on MMP-3 expression in PDL cells and the mechanism of this process were examined. Human PDL cells were exposed to bFGF at various concentrations (0.01-10 ng/ml) in monolayer cultures. bFGF increased [3H]thymidine incorporation and suppressed proteoglycan synthesis concentration-dependently. However, similar concentration ranges of bFGF increased the release of the cell-associated proteoglycans into the medium. Furthermore, bFGF increased MMP-3 mRNA levels concentration-dependently as examined by reverse transcription-polymerase chain reaction (RT-PCR). Induction of MMP-3 after the stimulation with bFGF was observed as early as 12 h with maximal at 24 h. Thereafter, the MMP-3 mRNA level gradually decreased until 72 h. Cycloheximide blocked the induction of MMP-3 by bFGF, indicating the requirement of de novo protein synthesis for this stimulation. Furthermore, MMP-3 expression induced by bFGF was abrogated by U0126, a specific inhibitor of MEK1/2 and ERK1/2 in mitogen-activated protein (MAP) kinase pathway, not by PD98059, a specific inhibitor of MEK1. In addition, bFGF up-regulated the phosphorylated ERK1/2 in 5 min with the maximal at 20 min as examined by Western blotting, and U0126 inhibited the ERK1/2 phosphorylation induced by bFGF. These findings suggest that bFGF induces MMP-3 expression in PDL cells through the activation of the MEK2 in MAP kinase pathway. bFGF stimulation on MMP-3 synthesis may be involved in the control of the cell-associated proteoglycans in PDL cells during periodontal regeneration and degradation.
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PMID:Basic fibroblast growth factor induces the expression of matrix metalloproteinase-3 in human periodontal ligament cells through the MEK2 mitogen-activated protein kinase pathway. 1260 5

We have previously reported that prostaglandin F2 alpha (PGF2 alpha) activates p44/p42 mitogen-activated protein kinase (MAPK) through protein kinase C (PKC) in osteoblast-like MC3T3-E1 cells. In the present study, we investigated the mechanism of vascular endothelial growth factor (VEGF) synthesis induced by PGF2 alpha and the effect of incadronate on the VEGF synthesis in these cells. PGF2 alpha significantly stimulated the VEGF synthesis in a dose-dependent manner between 1 pm and 10 microm. Cycloheximide reduced the PGF2 alpha effect. PGF2 alpha increased the levels of mRNA for VEGF. Cloprostenol, a PGF2 alpha-sensitive receptor agonist, potently induced the VEGF synthesis. Indomethacin, an inhibitor of cyclooxygenase, significantly reduced the PGF2 alpha-induced VEGF synthesis. Bisindolylmaleimide, an inhibitor of PKC, reduced the PGF2 alpha-induced VEGF synthesis. The VEGF synthesis induced by PGF2 alpha was significantly attenuated in the PKC down-regulated cells. PGF2 alpha elicited the translocation of PKC beta I from cytosol to membrane fraction. PD98059 or U0126, inhibitors of MEK, suppressed the VEGF synthesis induced by PGF2 alpha. Farnesyltransferase inhibitor failed to affect the PGF2 alpha-induced VEGF synthesis. Incadronate enhanced the synthesis of VEGF induced by PGF2 alpha. NaF-induced VEGF synthesis was also amplified by incadronate. PD98059 suppressed the enhancement by incadronate of PGF2 alpha-induced VEGF synthesis. Incadronate markedly enhanced the phosphorylation of Raf-1, MEK1/2, and p44/p42 MAPK induced by PGF2 alpha or 12-O-tetradecanoylphorbol-13-acetate, a PKC activator. Incadronate significantly enhanced the cloprostenol-increased level of VEGF concentration in mouse plasma in vivo. These results strongly suggest that PGF2 alpha stimulates VEGF synthesis through the PKC-dependent activation of p44/p42 MAPK in osteoblasts and that the incadronate enhances the VEGF synthesis at the point between PKC and Raf-1.
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PMID:Incadronate amplifies prostaglandin F2 alpha-induced vascular endothelial growth factor synthesis in osteoblasts. Enhancement of MAPK activity. 1264 77

Using cultured rat alveolar NR 8383 macrophages, this study investigated the effect of YC-1 [3-(5'-hydroxymethyl-2'-furyl)-1-benzyl indazole], a soluble guanylyl cyclase (sGC) activator, on the production of tumor necrosis factor-alpha (TNF alpha). YC-1 enhanced lipopolysaccharide and interferon-gamma (LPS/IFN gamma)-induced TNF alpha formation in a concentration- and time-dependent fashion. YC-1 also caused an increasing effect on the TNF alpha mRNA level, suggesting that the transcriptional process was involved. However, further studies suggested that cyclic GMP did not mediate the potentiation of YC-1 on TNF alpha release, because (a) the sGC inhibitor and the protein kinase G inhibitor failed to block the effect; and (b) the cyclic GMP analogues, on the contrary, concentration-dependently diminished LPS/IFN gamma-induced TNF alpha synthesis. In agreement with this finding, YC-1 produced changes in cell function but no changes in cyclic GMP and cyclic AMP levels or sGC activity. Pretreatment of the cells with cyclooxygenase inhibitors, a p38 mitogen-activated protein kinase inhibitor, a mitogen-activated protein kinase kinase (MEK) inhibitor, and a tyrosine kinase inhibitor did not attenuate the potentiation of TNF alpha release by YC-1. Cycloheximide prevented the YC-1-enhanced TNF alpha formation, implying that new protein synthesis was required. Interestingly, protein kinase C inhibitors enhanced the potentiation of YC-1 to a greater extent. Nevertheless, a protein kinase C activator, phorbol 12-myristate 13-acetate, failed to suppress the potentiation of TNFalpha production by YC-1. In summary, potentiation of TNF alpha release by YC-1 in LPS/IFN gamma-activated alveolar macrophages is an additional mode of action of this compound that is independent of the elevation of cyclic GMP. Thus, caution needs to be used in attributing the YC-1-mediated response to the activation of sGC.
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PMID:Potentiation of tumor necrosis factor-alpha expression by YC-1 in alveolar macrophages through a cyclic GMP-independent pathway. 1281 75

Mechanical strain plays a crucial role in bone remodeling during growth and development and healing of bone besides systemic and local factors. One of the major factors involves in remodeling process is matrix metalloproteinases (MMPs) such as MMP-13 that has been shown to degrade the native interstitial collagens in several tissues. To study how mechanical strain affects extracellular matrix degradation by MMP-13, a biaxial strain was applied to MC3T3-E1 osteoblastic cells plated onto a collagen-coated flexible elastic membrane. The MMP-13 protein and mRNA expression were determined by Western blotting and reverse transcriptase-PCR, respectively. The zymographic activities of MMP-13 increased dramatically at 30 min, reached a peak by 2-fold at 1 h, and maintained up to 4 h. Moreover, the MMP-13 and c-fos mRNA expressed at 5 min, increased to 2.8- and 3-fold at 1 h, respectively, and gradually declined thereafter. Cycloheximide and actinomycin D did not inhibit the MMP-13 and c-fos mRNA expression, suggesting that such expression does not require de novo protein synthesis and not change their stabilities. To investigate which of the mitogen-activated protein kinase (MAPK) pathways involves in the expression of MMP-13, inhibitors such as PD98059, SB203580, and SP600125 were used. However, only PD98059 (an inhibitor of MEK1/2 activation) inhibited MMP-13 and c-fos gene expression; the result was further substantiated by transfecting with the dominant negative mutants of MEK1/2 (MEK K97R) and ERK2. Taken together, our results showed that mechanical strain induces the MMP-13 expression through MEK-ERK signaling pathway to regulate mechanical adaptation.
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PMID:Mechanical strain induces collagenase-3 (MMP-13) expression in MC3T3-E1 osteoblastic cells. 1504 66

The eggs of Urechis unicinctus Von Drasche, an echiuroid, are arrested at P-I stage in meiosis. The meiosis is reinitiated by fertilization. Immunoblotting analysis using anti-ERK2 and anti-phospho-MAPK antibodies revealed a 44 kDa MAP kinase species that was constantly expressed in U. unicinctus eggs, quickly phosphorylated after fertilization, and dephosphorylated slowly before the completion of meiosis I. Phosphorylation of the protein was not depressed by protein synthesis inhibitor Cycloheximide (CHX), but was depressed by the MEK1 inhibitor PD98059. Under PD98059 treatment, polar body extrusion was suppressed and the function of centrosome and spindle was abnormal though GVBD was not affected, indicating that MAP kinase cascade was important for meiotic division of U. unicinctus eggs. Other discovery includes: A23187 and OA could parthenogenetically activate U. unicinctus eggs and phosphorylated 44 kDa MAP kinase species, indicating that the effect of fertilization on reinitiating meiosis and phosphorylation of 44 kDa MAP kinase specie is mediated by raising intracellular free calcium and by phosphorylation of some proteins, and that phosphotase(s) sensitive to OA is responsible for arresting U. unicinctus eggs in prophase I. diC8, an activator of PKC, accelerated the process of U. unicinctus egg meiotic division after fertilization and accelerated the dephosphorylation of 44 kDa MAP kinase specie, which implied that the acceleration effect of PKC on meiotic division was mediated by inactivation of MAP kinase cascade. Elevating cAMP/PKA level in U. unicinctus eggs had no effect on meiotic division of the eggs.
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PMID:Effects of MAP kinase pathway and other factors on meiosis of Urechis unicinctus eggs. 1573 26


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