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)

Desensitization of p21(ras) after stimulation of cells by growth factors and phorbol 12-myristate 13-acetate (PMA) correlates with hyperphosphorylation of the guanine nucleotide exchange factor Son-of-sevenless (Sos) and its dissociation from the adaptor protein Grb2 (Cherniack, A., Klarlund, J. K., Conway, B. R., and Czech, M. P. (1995) J. Biol. Chem. 270, 1485-1488). To test the role of the Raf/mitogen-activated protein (MAP) kinase pathway, we utilized cells expressing a chimera composed of the catalytic domain of p74Raf-1 and the hormone binding domain of the estradiol receptor (DeltaRaf-1:ER). Estradiol markedly stimulated DeltaRaf-1:ER and the downstream MEK and MAP kinases in these cells as well as Sos phosphorylation. However, the dissociation of Grb2 from Sos observed in response to PMA was not apparent upon DeltaRaf-1:ER activation. Furthermore, stimulation of DeltaRaf-1:ER did not impair GTP loading of p21(ras) in response to platelet-derived growth factor or epidermal growth factor. We conclude that activation of the Raf/MAP kinase pathway alone in these cells is insufficient to cause disassembly of Sos from Grb2 or to interrupt the ability of Sos to catalyze activation of p21(ras).
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PMID:Role of the Raf/mitogen-activated protein kinase pathway in p21ras desensitization. 866 95

To understand the molecular mechanism by which the angiotensin II (AII) type 1 receptor (AT1 receptor) transduces its biological signal, we examined the role of various signaling molecules involved in AT1 receptor signaling in Chinese hamster ovary cells stably transfected with the AT1 receptor. AT1 receptor-transfected cells responded to AII treatment by inhibiting adenylyl cyclase, increasing the intracellular Ca2+ concentration, and activating protein kinase C (PKC) alpha and PKC epsilon. AII also activated the c-fos gene and mitogen-activated protein (MAP) kinases. The activation of PKC, the c-fos gene, and MAP kinases was blocked by inhibition of PKC induced by pretreatment with 12-O-tetradecanoylphorbol-13-acetate but not by pretreatment with pertussis toxin, suggesting that PKC couples to the activation of the the c-fos gene and MAP kinases. In addition, AII activated Raf-1 and MAP kinase kinase in a PKC-dependent manner. A dominant negative mutant of Ras had no effect on AII-induced MAP kinase or c-fos gene activation. Thus, the AT1 receptor signals through Raf-1 and its downstream signaling molecules by a PKC-dependent mechanism that does not involve Ras activation.
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PMID:Angiotensin II type 1 receptor signals through Raf-1 by a protein kinase C-dependent, Ras-independent mechanism. 879 90

We reported recently that angiotensin II (AII) and phorbol 12-myristate 13-acetate (PMA) transiently inhibit interleukin 6 (IL-6)-stimulated tyrosine phosphorylation of signal transducers and activators of transcription 3 (Stat3) and subsequent formation of sis-inducing factor-A (SIF-A). However, the AII-mediated inhibition was independent of PMA-sensitive isoforms of protein kinase C (Bhat, G. J., Thekkumkara, T. J., Thomas, W. G., Conrad, K. M., and Baker, K. M. (1995) J. Biol. Chem. 270, 19059-19065). In this study, we demonstrate that the inhibition of IL-6-induced Stat3/SIF-A by AII is concentration-dependent and does not involve degradation of Stat3 protein. We hypothesized that the activation profile of the AII- and PMA-induced mitogen-activated protein (MAP) kinase cascade may be different from that of IL-6 and could contribute to the inhibitory effect; therefore, blocking the MAP kinase pathway at the level of MAPK kinase (MAPKK) would attenuate this inhibitory effect. AII and PMA rapidly induced high levels of MAP kinase activity (8-fold), which contrasted with the delayed and weak activation by IL-6 (1. 7-fold). Treatment of cells with PD98059, a specific inhibitor of MAPKK1, attenuated the inhibitory effects of AII and PMA on IL-6-induced Stat3 tyrosine phosphorylation and SIF-A formation. These data suggest that differences in magnitude and/or duration of activation of the MAP kinase cascade differentially affects the status of Stat3 tyrosine phosphorylation, and that MAPKK1 or a downstream intermediate is involved in the inhibition of IL-6-induced Stat3 by AII and PMA. Modulatory cross-talk between AII and IL-6 may have relevance in pathophysiological conditions such as cardiac hypertrophy and in acute phase and inflammatory responses.
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PMID:Angiotensin II interferes with interleukin 6-induced Stat3 signaling by a pathway involving mitogen-activated protein kinase kinase 1. 879 9

The serine/threonine kinase Raf-1 functions downstream of Rats in a signal transduction cascade which transmits mitogenic stimuli from the plasma membrane to the nucleus. Raf-1 integrates signals coming from extracellular factors and, in turn, activates its substrate, MEK kinase. MEK activates mitogen-activated protein kinase (MAPK), which phosphorylates other kinases as well as transcription factors. Raf-1 exists in a complex with HSP90 and other proteins. The benzoquinone ansamycin geldanamycin (GA) binds to HSP90 and disrupts the Raf-1-HSP90 multimolecular complex, leading to destabilization of Raf-1. In this study, we examined whether Raf-1 destabilization is sufficient to block the Raf-1-MEK-MAPK signalling pathway and whether GA specifically inactivates the Raf-1 component of this pathway. Using the model system of NIH 3T3 cells stimulated with phorbol 12-myristate 13-acetate (PMA), we show that GA does not affect the ability of protein kinase C alpha to be activated by phorbol esters, but it does block activation of MEK and MAPK. Further, GA does not decrease the activity of constitutively active MEK in transiently transfected cells. Finally, disruption of the Raf-1-MEK-MAPK signalling pathway by GA prevents both the PMA-induced proliferative response and PMA-induced activation of a MAPK-sensitive nuclear transcription factor. Thus, we demonstrate that interaction between HSP90 and Raf-1 is a sine qua non for Raf stability and function as a signal transducer and that the effects observed cannot be attributed to a general impairment of protein kinase function.
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PMID:Destabilization of Raf-1 by geldanamycin leads to disruption of the Raf-1-MEK-mitogen-activated protein kinase signalling pathway. 881 98

The purpose of this investigation was to pharmacologically probe the signaling pathways thought to be involved in protein kinase C (PKC)-stimulated superoxide anion (O2-) generation in all-trans retinoic acid-treated human promyelocytic HL-60 cell line (HL-60), targeting PKC, mitogen-activated protein kinase (MAPK), MAPK kinase (MEK), protein serine-threonine phosphatase(s) (PSP), protein tyrosine kinase(s) (PTK) and phosphatase(s) (PTP), secretory phospholipase A2, cyclooxygenase (CO) and 5-lipoxygenase with selected inhibitors. The following agents inhibited phorbol 12-myristate 13-acetate-stimulated O2- generation significantly in the all-trans retinoic acid-treated HL-60 cells (expressed as percentage of control, P < .05): 1) PKC inhibitors: staurosporine (100 nM, 3 +/- 1%); Ro 31-8220 (1 microM, 3 +/- 2%); sphingosine (100 microM, 15 +/- 7%); 2) PSP 1 and 2a inhibitors, okadaic acid (10 microM, 35 +/- 1%); calyculin A (10 microM, 73 +/- 1%); 3) MAPK inhibitor: SB-203580 (100 microM, 62 +/- 1%); 4) PTP inhibitors: phenylarsine oxide (1 microM, 12 +/- 9%); diamide (1 mM, 21 +/- 11%); and 5) secretory phospholipase A2 inhibitors: manoalide (1 microM, 24 +/- 10%); scalaradial (1 microM, 11 +/- 4%). Exogenously added arachidonic acid-stimulated O2- generation in a time- and dose-dependent manner. The following inhibitors enhanced or did not significantly affect phorbol 12-myristate 13-acetate-stimulated O2- generation (expressed as percentage of control): 1) PTK inhibitors: genistein (100 microM, 69 +/- 12%); CGP 53716 (100 microM, 67 +/- 10%); herbimycin A (10 microM, 67.4 +/- 1%); 2) PSP 2b inhibitors: cyclosporin A (30 microM, 71 +/- 5%); FK506 (30 microM, 88 +/- 7%); 3) CO inhibitor: indomethacin (100 microM, 111 +/- 12%); 4) 5-lipoxygenase inhibitor: WY 50,295 (100 microM, 140 +/- 23%); 5) MEK inhibitor: PD98059 (100 microM, 94 +/- 6.7%); and 6) the PTP inhibitor: orthovanadate (100 microM, 131 +/- 25%). Our pharmacological study suggests that, in neutrophil-like HL-60 cells, the signaling pathways leading to PMA-stimulated O2- generation appear to involve PKC, MAPK, phospholipase A2, arachidonic acid, PSP 1 and 2a and PTP. Furthermore, PTK, MEK, CO, 5-lipoxygenase and PSP 2b do not appear to participate in the modulation of PKC-stimulated O2- generation.
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PMID:Pharmacological targeting of signaling pathways in protein kinase C-stimulated superoxide generation in neutrophil-like HL-60 cells: effect of phorbol ester, arachidonic acid and inhibitors of kinase(s), phosphatase(s) and phospholipase A2. 893 Jan 66

Recently we have identified a mitogen-activated protein kinase (MAPK)-activated protein kinase, named 3pK (G. Sithanandam, F. Latif, U. Smola, R. A. Bernal, F.-M. Duh, H. Li, I. Kuzmin, V. Wixler, L. Geil, S. Shresta, P. A. Lloyd, S. Bader, Y. Sekido, K. D. Tartof, V. I. Kashuba, E. R. Zabarovsky, M. Dean, G. Klein, B. Zbar, M. I. Lerman, J. D. Minna, U. R. Rapp, and A. Allikmets, Mol. Cell. Biol. 16:868-876, 1996). In vitro characterization of the kinase revealed that 3pK is activated by ERK. It was further shown that 3pK is phosphorylated in vivo after stimulation of cells with serum. However, the in vivo relevance of this observation in terms of involvement of the Raf/MEK/ERK cascade has not been established. Here we show that 3pK is activated in vivo by the growth inducers serum and tetradecanoyl phorbol acetate in promyelocytic HL60 cells and transiently transfected embryonic kidney 293 cells. Activation of 3pK was Raf dependent and was mediated by the Raf/MEK/ERK kinase cascade. 3pK was also shown to be activated after stress stimulation of cells. In vitro studies with recombinant proteins demonstrate that in addition to ERK, members of other subgroups of the MAPK family, namely, p38RK and Jun-N-terminal kinases/stress-activated protein kinases, were also able to phosphorylate and activate 3pK. Cotransfection experiments as well as the use of a specific inhibitor of p38RK showed that these in vitro upstream activators also function in vivo, identifying 3pK as the first kinase to be activated through all three MAPK cascades. Thus, 3pK is a novel convergence point of different MAPK pathways and could function as an integrative element of signaling in both mitogen and stress responses.
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PMID:3pK, a novel mitogen-activated protein (MAP) kinase-activated protein kinase, is targeted by three MAP kinase pathways. 894 23

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

The mechanism of mitogen-activated protein kinase (MAPK, ERK) stimulation by the GnRH analog [D-Trp6]GnRH (GnRH-a) was investigated in the gonadotroph-derived alphaT3-1 cell line. GnRH-a as well as the protein kinase C (PKC) activator 12-O-tetradecanoyl phorbol-13-acetate (TPA) stimulated a sustained response of MAPK activity, whereas epidermal growth factor (EGF) stimulated a transient response. MAPK kinase (MEK) is also activated by GnRH-a, but in a transient manner. GnRH-a and TPA apparently activated mainly the MAPK isoform ERK1, as revealed by Mono-Q fast protein liquid chromatography followed by Western blotting as well as by gel kinase assay. GnRH-a and TPA stimulated the tyrosine phosphorylation of several proteins, and this effect as well as the stimulation of MAPK activity were inhibited by the PKC inhibitor GF 109203X. Similarly, down-regulation of TPA-sensitive PKC subspecies nearly abolished the effect of GnRH-a and TPA on MAPK activity. Furthermore, the protein tyrosine kinase (PTK) inhibitor genistein inhibited protein tyrosine phosphorylation and reduced GnRH-a-stimulated MAPK activity by 50%, suggesting the participation of genistein-sensitive and insensitive pathways in GnRH-a action. Although Ca2+ ionophores have only a marginal stimulatory effect, the removal of Ca2+ markedly reduced MAPK activation by GnRH-a and TPA, but had no effect on GnRH-a and TPA stimulation of protein tyrosine phosphorylation. Interestingly, the removal of Ca2+ also partly inhibited the activation of MAPK by EGF and vanadate/H2O2. Thus, a calcium-dependent component(s) downstream of PKC and PTK might also participate in MAPK activation. Elevation of cAMP by forskolin exerted partial inhibition on EGF, but not on TPA or GnRH-a action, suggesting that MEK activators other than Raf-1 might be involved in GnRH action. We conclude that Ca2+, PTK, and PKC participate in the activation of MAPK by GnRH-a, with Ca2+ being necessary downstream to PKC and PTK.
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PMID:Mechanism of mitogen-activated protein kinase activation by gonadotropin-releasing hormone in the pituitary of alphaT3-1 cell line: differential roles of calcium and protein kinase C. 907 30

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

We have developed a novel expression screening method for identifying protein kinase substrates. In this method, a lambda phage cDNA expression library is screened by in situ, solid-phase phosphorylation using purified protein kinase and [gamma-32P]ATP. Screening a HeLa cDNA library with ERK1 MAP kinase yielded cDNAs of previously characterized ERK substrates, c-Myc and p90RSK, demonstrating the utility of this method for identifying physiological protein kinase substrates. A novel clone isolated in this screen, designated MNK1, encodes a protein-serine/threonine kinase, which is most similar to MAP kinase-activated protein kinase 2 (MAPKAP-K2), 3pK/MAPKAP-K3 and p90RSK. Bacterially expressed MNK1 was phosphorylated and activated in vitro by ERK1 and p38 MAP kinases but not by JNK/SAPK. Further, MNK1 was activated upon stimulation of HeLa cells with 12-O-tetradecanoylphorbol-13-acetate, fetal calf serum, anisomycin, UV irradiation, tumor necrosis factor-alpha, interleukin-1beta, or osmotic shock, and the activation by these stimuli was differentially inhibited by the MEK inhibitor PD098059 or the p38 MAP kinase inhibitor SB202190. Together, these results indicate that MNK1 is a novel class of protein kinase that is activated through both the ERK and p38 MAP kinase signaling pathways.
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PMID:MNK1, a new MAP kinase-activated protein kinase, isolated by a novel expression screening method for identifying protein kinase substrates. 915 18


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