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)

Both angiotensin II (Ang II) and platelet-derived growth factor (PDGF) rapidly increase intracellular Ca2+ and activate protein kinase C (PKC) and MAP kinase in vascular smooth muscle cells (VSMCs). However, Ang II causes cell hypertrophy, whereas PDGF causes hyperplasia. These findings indicate that VSMCs are a good model for studying the relationship between cell growth and the MAP kinase pathway. In this study, we investigated the role of Raf in activation of 42- and 44-kD MAP kinases. Western blot analysis showed that c-Raf-1 was the predominant Raf isozyme in cultured rat aortic VSMCs. In response to Ang II, there was translocation of Raf to the membrane, which occurred significantly earlier than MAP kinase activation, suggesting that Raf activation precedes MAP kinase activation. Translocation of Raf to the membrane resulted in association with H-Ras as shown by c-Raf-1 coprecipitation with anti-Ras anti-bodies. Western blot analysis of H-Ras immunoprecipitates revealed c-Raf-1, but c-mos, MEK (MAP kinase/extracellular signal-regulated kinase) kinase-1 (MEKK-1), and Raf-B were not present. MAP kinase kinase kinase (MAPKKK) activity was assayed in c-Raf-1 and H-Ras immunoprecipitates by MAP kinase kinase-dependent phosphorylation of catalytically inactive 42-kD MAP kinase. In Ras immunoprecipitates, MAPKKK activity was stimulated approximately threefold by both Ang II and PDGF, with a peak at 5 minutes. Downregulation of PKC by 24-hour exposure to phorbol ester significantly inhibited Ang II-stimulated and PDGF-stimulated MAPKKK activity (approximately 80% decrease) and Raf translocation (approximately 90% decrease), suggesting that a phorbol-responsive PKC is upstream from MAPKKK and Raf. In contrast, Ang II (but not PDGF) stimulation of MAP kinase was unaffected by PKC downregulation or pharmacological PKC inhibition. These findings demonstrate for the first time that Ang II stimulation of MAP kinase may occur via a pathway independent of c-Raf-1 and of the phorbol-responsive PKC isozymes. The differing effects of Ang II and PDGF on VSMC growth may be a consequence of specific signal transduction events, as demonstrated here for activation of MAP kinase.
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PMID:Angiotensin II stimulates MAP kinase kinase kinase activity in vascular smooth muscle cells, Role of Raf. 888 93

We have investigated the requirement for mitogen-activated protein (MAP) kinase in the stimulation of DNA synthesis by platelet-derived growth factor (PDGF) in rat aortic smooth muscle cells using a phosphorothioate-modified oligodeoxy-nucleotide (ODN) to deplete MAP kinase. Treatment for 72 h with MAP kinase antisense ODN directed against both the p42 and p44 isoforms of MAP kinase abolished the expression of MAP kinase and reduced agonist-stimulated MAP kinase activity by approx. 95%. The scrambled control ODN was without effect, but the sense control ODN slightly enhanced the expression of both isoforms. Abolition of MAP kinase activity by antisense ODN treatment prevented angiotensin II- and PDGF-stimulated activation of p90 ribosomal S6 kinase activity, but did not affect activation of MAP kinase kinase. In addition, antisense ODN pretreatment reduced PDGF-stimulated [3H]thymidine incorporation to < 5% of control, and decreased basal incorporation by approx. 90%. In contrast, basal [3H]thymidine incorporation was enhanced approx. 60% by control sense ODN treatment. These results indicate an obligatory role for MAP kinase in the activation of a number of early events in mitogenesis, including DNA synthesis, in vascular smooth muscle cells.
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PMID:Treatment of vascular smooth muscle cells with antisense phosphorothioate oligodeoxynucleotides directed against p42 and p44 mitogen-activated protein kinases abolishes DNA synthesis in response to platelet-derived growth factor. 894 76

In order to elucidate the signal transduction pathway from external mechanical stress to nuclear gene expression in mechanical stress-induced cardiac hypertrophy, we examined the time course of activation of Raf-1 kinase (Raf-1), mitogen-activated protein kinase kinase (MAPKK) and MAP kinases (MAPKs) in neonatal rat cardiac myocytes. Mechanical stretch transiently activated Raf-1 and MAPKK with a peak at 2 and 5 min after stretch, respectively. In addition, MAPKs were maximally activated at 8 min after stretch. Next, the relationship between stretch-induced hypertrophy and the cardiac reninangiotensin system was investigated. When the stretch-conditioned culture medium was transferred to non-stretched cardiac myocytes, the medium activated MAPK activity slightly but significantly, and the activation was completely blocked by the type I angiotensin II (AngII) receptor antagonist, CV-11974. Moreover, in in vivo studies using spontaneously hypertensive rats, hypertension-induced cardiac hypertrophy was significantly reduced by treatment with subpressure doses of CV-11974. In addition, CV-11974 reduced the isozymic transition of MHC from VI to V3 and inhibited the accumulation of collagen fibers in the extracellular space of the myocardium. These results suggest that mechanical stress activates the protein kinase cascade of phosphorylation in cardiac myocytes in the order of Raf-1, MAPKK and MAPKs. AngII, which is secreted from stretched myocytes, possibly activates these protein kinases. Moreover, it was shown that CV-11974 causes regression of cardiac hypertrophy and has cardioprotective effects on hypertrophied myocardium in vivo.
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PMID:Angiotensin II mediates mechanical stress-induced cardiac hypertrophy. 896 84

To elucidate the signal transduction pathway from external stimuli to nuclear gene expression in mechanical stress-induced cardiac hypertrophy, we examined the time course of activation of protein kinases such as Raf-1 kinase (Raf-1), mitogen-activated protein kinase kinase (MAPKK), MAP kinases (MAPKs) and 90-kDa ribosomal S6 kinase (p90rsk) in neonatal rat cardiomyocytes. Mechanical stretch rapidly activated Raf-1 and its maximal activation was observed at 1-2 min after stretch. The activity of MAPKK was also increased by stretch, with a peak at 5 min after stretch. In addition, MAPKs and p90rsk were maximally activated at 8 min and at 10-30 min after stretch, respectively. Next, the relationship between mechanical stress-induced hypertrophy and the cardiac renin-angiotensin system was investigated. When the stretch-conditioned culture medium was transferred to the culture dish of non-stretched cardiac myocytes, the medium activated MAPK activity slightly but significantly, and the activation was completely blocked by the type 1 angiotensin II receptor antagonist, CV-11974. However, activation of Raf-1 and MAPKs provoked by stretching cardiomyocytes was only partially suppressed by pretreatment with CV-11974. These results suggest that mechanical stress activates the protein kinase cascade of phosphorylation in cardiac myocytes in the order of Raf-1, MAPKK, MAPKs and p90rsk, and that angiotensin II, which is secreted from stretched myocytes, activates a part of these protein kinases.
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PMID:Molecular aspects of mechanical stress-induced cardiac hypertrophy. 897 57

To investigate the molecular basis of the hypertrophic action of angiotensin II (AII) in vascular smooth muscle cells (SMC), we have examined the ability of the hormone to regulate the function of the translational repressor 4E-binding protein 1 (4E-BP1). Addition of AII to quiescent aortic SMC potently increased the phosphorylation of 4E-BP1 as revealed by a decreased electrophoretic mobility and an increased phosphate content of the protein. The stimulation of 4E-BP1 phosphorylation was maximal at 15 min and persisted up to 120 min. Results from affinity chromatography on m7GTP-agarose demonstrated that AII-induced phosphorylation of 4E-BP1 promotes its dissociation from eIF4E in target cells. Further characterization of 4E-BP1 phosphorylation by phosphoamino acid analysis and phosphopeptide mapping revealed that 4E-BP1 is phosphorylated on eight distinct peptides containing serine and threonine residues in AII-treated cells. The combination of results obtained from kinetics experiments, phosphopeptide analysis of in vitro and in vivo phosphorylated 4E-BP1, and pharmacological studies with the MAP kinase kinase inhibitor PD 98059 provided strong evidence that the MAP kinases ERK1/ERK2 are not involved in the regulation of 4E-BP1 phosphorylation in aortic SMC. Together, our results demonstrate that AII treatment of vascular SMC leads to hyperphosphorylation of the translational regulator 4E-BP1 and to its dissociation from eIF4E by a MAP kinase-independent mechanism.
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PMID:Angiotensin II stimulates phosphorylation of the translational repressor 4E-binding protein 1 by a mitogen-activated protein kinase-independent mechanism. 902 Jan 7

Activation of 44 and 42 kDa extracellular signal-regulated kinases (ERK)1/2 by angiotensin II (angII) plays an important role in vascular smooth muscle cell (VSMC) function. The dual specificity mitogen-actived protein (MAP) kinase/ERK kinase (MEK) activates ERK1/2 in response to angII, but the MEK activating kinases remain undefined. Raf is a candidate MEK kinase. However, a kinase other than Raf appears responsible for angII-mediated signal transduction because we showed previously that treatment with 1 microM phorbol 12, 13-dibutyrate (PDBU) for 24 h completely blocked Raf-Ras association in VSMC but did not inhibit activation of MEK and ERK1/2 by angII. We hypothesized that an atypical protein kinase C (PKC) isoform, which lacks a phorbol ester binding domain, mediated ERK1/2 activation by angII. Western blot analysis of rat aortic VSMC with PKC isoform-specific antibodies showed PKC-alpha, -beta1, -delta, -epsilon, and -zeta in relative abundance. All isoforms except PKC-zeta were down-regulated by 1 microM PDBU for 24 h suggesting that PKC-zeta was responsible for angII-mediated ERK1/2 activation. In response to angII, PKC-zeta associated with Ras as shown by co-precipitation of PKC-zeta with anti-H-Ras antibody. To characterize further the role of PKC-zeta, PKC-zeta protein was depleted specifically by transfection with antisense PKC-zeta oligonucleotides. Antisense PKC-zeta oligonucleotide treatment significantly decreased PKC-zeta protein expression (without effect on other PKC isoforms) and angII-mediated ERK1/2 activation in a concentration-dependent manner. In contrast, ERK1/2 activation by platelet-derived growth factor and phorbol ester was not significantly inhibited. These results demonstrate an important difference in signal transduction by angII compared with PDGF and phorbol ester in VSMC, and suggest a critical role for PKC-zeta and Ras in angII stimulation of ERK1/2.
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PMID:Protein kinase C-zeta mediates angiotensin II activation of ERK1/2 in vascular smooth muscle cells. 904 26

In rat aortic smooth muscle cells (RASMC), pretreatment with forskolin inhibited the activation of p42/44 isoforms of mitogen-activated protein kinase (MAP) kinase stimulated in response to low concentrations of PDGF (10 ng/ml). This correlated with a strong inhibition of PDGF-stimulated MEK and C-Raf-1 kinase activity. However, the effect of forskolin could be surmounted by increasing the concentration of PDGF. Under such conditions forskolin was only effective against prolonged MAP kinase activation. The ability of forskolin to inhibit the late phase of MAP kinase activity was reversed by pretreatment of the cells with cycloheximide, suggesting the involvement of a protein synthesis step. This was not due to effects upstream of MAP kinase since PDGF-stimulated MEK activation was decreased by cycloheximide, an effect potentiated by forskolin. Forskolin stimulated the induction of the dual specific phosphatase MAP kinase phosphatase-1 (MKP-1), although this effect was small relative to levels induced by PDGF and angiotensin II. However, PDGF stimulated induction of MKP-1 was abolished by the protein kinase A inhibitor H89 and this correlated with the reversal of forskolin-mediated inhibition of PDGF-stimulated MAP kinase activity. These studies implicate a role for intracellular cyclic AMP in at least two aspects of MAP kinase signaling, including both the inhibition of Raf-1 activation and the induction of MKP-1.
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PMID:Cyclic AMP inhibitors inhibits PDGF-stimulated mitogen-activated protein kinase activity in rat aortic smooth muscle cells via inactivation of c-Raf-1 kinase and induction of MAP kinase phosphatase-1. 921 35

We examined the signal transduction pathway for the development of cardiac hypertrophy induced by high blood pressure. The activities of Raf-1 kinase (Raf-1), mitogen-activated protein kinase kinase (MAPKK), MAP kinases (MAPKs) and 90-kDa ribosomal S6 kinase (p90rsk) was examined by passively stretching neonatal rat cardiomyocytes in vitro. Mechanical stretch activated these protein kinases transiently and sequentially: the maximal activation of Raf-1, MAPKK, MAPKs and p90rsk was observed at 2 minutes, 5 minutes, 8 minutes and 10 approximately 30 minutes, respectively. Both angiotensin II (AngII) and endothelin-1 (ET-1) were constitutively secreted from cultured cardiomyocytes, and a significant increase in the concentration was recognized in the culture medium of cardiomyocytes within 10 minutes after stretch. ET-1 mRNA levels were also increased in cardiomyocytes at 30 minutes after stretch. Moreover, ET-1 and AngII synergistically activated Raf-1 and MAPKs in cultured cardiomyocytes. In conclusion, mechanical stretch stimulates secretion and production of AngII and ET-1 in cultured cardiomyocytes, and both vasoconstrictive peptides may play an important role in mechanical stress (high blood pressure)-induced cardiac hypertrophy.
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PMID:[Molecular mechanism of cardiac hypertrophy and dysfunction]. 928 12

In vascular smooth muscle cells, the induction of early growth response genes involves the Janus kinase (JAK)/signal transducer and activators of transcription (STAT) and the Ras/Raf-1/mitogen-activated protein kinase cascades. In the present study, we found that electroporation of antibodies against MEK1 or ERK1 abolished vascular smooth muscle cell proliferation in response to either platelet-derived growth factor or angiotensin II. However, anti-STAT1 or -STAT3 antibody electroporation abolished proliferative responses only to angiotensin II and not to platelet-derived growth factor. AG-490, a specific inhibitor of the JAK2 tyrosine kinase, prevented proliferation of vascular smooth muscle cells, complex formation between JAK2 and Raf-1, the tyrosine phosphorylation of Raf-1, and the activation of ERK1 in response to either angiotensin II or platelet-derived growth factor. However, AG-490 had no effect on angiotensin II- or platelet-derived growth factor-induced Ras/Raf-1 complex formation. Our results indicate that: 1) STAT proteins play an essential role in angiotensin II-induced vascular smooth muscle cell proliferation, 2) JAK2 plays an essential role in the tyrosine phosphorylation of Raf-1, and 3) convergent mitogenic signaling cascades involving the cytosolic kinases JAK2, MEK1, and ERK1 mediate vascular smooth muscle cell proliferation in response to both growth factor and G protein-coupled receptors.
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PMID:Role of Janus kinase/signal transducer and activator of transcription and mitogen-activated protein kinase cascades in angiotensin II- and platelet-derived growth factor-induced vascular smooth muscle cell proliferation. 930 39

The neuronal angiotensin II (Ang II) type 1 (AT1) receptor is coupled to the Ras-Raf-1-mitogen-activated protein (MAP) kinase signal-transduction pathway (Yang H, Lu D, Yu K, Raizada MK. Regulation of neuromodulatory actions of angiotensin II in the brain neurons by the Ras-dependent mitogen-activated protein kinase pathway. J Neurosci. 1996;16:4047-4058). In this study we compared the effects of angiotensin II (Ang II) on AT1 receptor phosphorylation and the ability of the phosphorylated receptor to bind Ang II in neuronal cultures of Wistar-Kyoto rat (WKY) and spontaneously hypertensive rat (SHR) brains to further our understanding of the Ang II signaling mechanism. Ang II caused a time-dependent phosphorylation of AT1 receptors in both WKY and SHR brain neurons. The level of phosphorylation was higher in the SHR brain neurons; this finding was consistent with increased AT1 receptors in these cells. MAP kinase was involved in this phosphorylation, a conclusion supported by the following evidence: (1) exogenous MAP kinase phosphorylated the AT1 receptor; (2) PD98059, a MAP kinase kinase inhibitor, attenuated Ang II-stimulated AT1 receptor phosphorylation; and (3) MAP kinase and AT1 receptors were coimmunoprecipitated in Ang II-stimulated neurons. Finally, MAP kinase phosphorylation was associated with the loss of 125I-[Sar1-Ile8]-Ang II binding ability of the AT1 receptor in both strains of neurons. These observations show that Ang II stimulates phosphorylation of the neuronal AT1 receptor by a mechanism involving MAP kinase and that the phosphorylated neuronal AT1 receptor does not exhibit Ang II binding activity in the brains of either WKY or SHR.
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PMID:Angiotensin II-induced phosphorylation of the AT1 receptor from rat brain neurons. 931 16


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