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)

Insulin stimulates the Ras/Raf/MEK/ERK pathway leading to feedback phosphorylation of the Ras guanylnucleotide exchange protein SOS and dissociation of Grb2 from SOS. Even though epidermal growth factor (EGF) also stimulates ERK activity and phosphorylation of SOS similar to insulin, EGF induces a dissociation of the Grb2-SOS complex from Shc. To determine the molecular basis for this difference, we examined the signaling properties of a mutant EGF receptor lacking the five major autophosphorylation sites. Although EGF stimulation of the mutant EGF receptor activates ERK and phosphorylation of both Shc and SOS, it fails to directly associate with either Shc or Grb2. However, under these conditions EGF induces a dissociation of the Grb2-SOS complex suggesting a role for receptor and/or plasma membrane targeting in the stabilization of Grb2-SOS interaction. Consistent with this hypothesis, expression of an SH2 domain Grb2 mutant which is unable to mediate plasma membrane targeting of the Grb2-SOS complex results in both insulin- and EGF-stimulated uncoupling of Grb2 from SOS. Furthermore, a plasma membrane-bound Grb2 fusion protein remains constitutively associated with SOS. Together, these data demonstrate that EGF stimulation prevents the feedback uncoupling of Grb2 from SOS by inducing a persistent plasma membrane receptor targeting of the Grb2-SOS complex.
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PMID:Epidermal growth factor receptor targeting prevents uncoupling of the Grb2-SOS complex. 862 25

The mitogen-activated protein kinase (MAPK) family is comprised of key regulatory proteins that control the cellular response to both proliferation and stress signals. In this study we investigated the factors controlling MAPK activation by H2O2 and explored the impact of altering the pathways to kinase activation on cell survival following H2O2 exposure. Potent activation (10-20-fold) of extracellular signal-regulated protein kinase (ERK2) occurred within 10 min of H2O2 treatment, whereupon rapid inactivation ensued. H2O2 activated ERK2 in several cell types and also moderately activated (3-5-fold) both c-Jun N-terminal kinase and p38/RK/CSBP. Additionally, H2O2 increased the mRNA expression of MAPK-dependent genes c-jun, c-fos, and MAPK phosphatase-1. Suramin pretreatment completely inhibited H2O2 stimulation of ERK2, highlighting a role for growth factor receptors in this activation. Further, ERK2 activation by H2O2 was blocked by pretreatment with either N-acetyl-cysteine, o-phenanthroline, or mannitol, indicating that metal-catalyzed free radical formation mediates the initiation of signal transduction by H2O2. H2O2-stimulated activation of ERK2 was abolished in PC12 cells by inducible or constitutive expression of the dominant negative Ras-N-17 allele. Interestingly, PC12/Ras-N-17 cells were more sensitive than wild-type PC12 cells to H2O2 toxicity. Moreover, NIH 3T3 cells expressing constitutively active MAPK kinase (MEK, the immediate upstream regulator of ERK) were more resistant to H2O2 toxicity, while those expressing kinase-defective MEK were more sensitive, than cells expressing wild-type MEK. Taken together, these studies provide insight into mechanisms of MAPK regulation by H2O2 and suggest that ERK plays a critical role in cell survival following oxidant injury.
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PMID:Activation of mitogen-activated protein kinase by H2O2. Role in cell survival following oxidant injury. 862 53

We report that recombinant glia maturation factor (GMF), a 17-kDa brain protein, inhibits the activity of mitogen-activated protein (MAP) kinase in the test tube assay, in particular the ERK1/ERK2 isoforms. A preliminary phosphorylation of GMF by protein kinase A (PKA) dramatically increases its inhibitory effect by over 600-fold (Ki approximately 3 nM), making it the most potent MAP kinase inhibitor ever reported. Immunoprecipitation of GMF from cell extracts using its specific antibody coprecipitates ERK (and vice versa), suggesting the association of the two proteins in the cell. The inhibitory effect of PKA-phosphorylated GMF is specific, as it does not suppress the activity of cdc2 kinase, another proline-directed kinase. Nor does it inhibit MAP kinase kinase (MEK) and MAP kinase-activated protein (MAPKAP) kinase-2, the two enzymes immediately upstream and downstream, respectively, of ERK. Of the other three enzymes that can phosphorylate GMF, only p90 ribosomal S6 kinase (RSK) enhances the inhibitory function of GMF on ERK; protein kinase C (PKC) and casein kinase II (CKII) are without effect. The inhibition of ERK by PKA-phosphorylated GMF suggests that GMF could be one of the mediators of the suppressive effect of the PKA pathway on the MAP kinase pathway. On the other hand, that RSK-phosphorylated GMF also inhibits ERK implies a negative feedback loop in the regulation of MAP kinase activity.
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PMID:In vitro inhibition of MAP kinase (ERK1/ERK2) activity by phosphorylated glia maturation factor (GMF). 863 70

Age-related changes in the functional properties of human T cells are well described, but less is known about possible changes in T cell signaling pathways. The signaling pathways mediated by mitogen-activated protein kinases (MAPK) are considered essential for normal cellular growth and function. Several stimuli trigger MAPK activation in human T cells and MEK (MAPK or ERK kinases) are immediate upstream inducers of MAPK activation. The current study investigated if aging might influence the activation and expression of MAPK and MEK in human T cells. Exposure of peripheral blood T cells from young subjects to PHA or cross-linked anti-CD3 monoclonal antibodies stimulated rapid increases in MAPK and MEK enzymatic activity. By contrast, significant reductions of MAPK and MEK activation were observed in stimulated T cells from 7 of 13 elderly subjects. Kinetic studies showed that the age-related impairments represented reduction in both the levels and duration of MAPK activation. In addition, Western immunoblot analysis did not reveal significant age-related differences in T cell expression of p42mapk/ERK2, p44mapk/ERK1, or MEK, suggesting impairments in upstream inducers of MEK/MAPK activation. Other experiments determined if agents that directly stimulate upstream Ras or Raf kinase components of the early MAPK cascade might reverse the age-related impairments of MAPK activation. Treatment of elderly T cells with fluoroaluminate (AlF(-)4), phorbol esters/Ca2+ ionophores, or okadaic acid stimulated increased MAPK activation compared to anti-CD3. However, these agents failed to restore MAPK activation in elderly T cells to the levels seen in young T cells. These results suggest that aberrancies in the MAPK activation cascade may underlie the age-related reductions of MAPK activation in human T cells stimulated via the TCR/CD3 complex.
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PMID:Age-related reductions in the activation of mitogen-activated protein kinases p44mapk/ERK1 and p42mapk/ERK2 in human T cells stimulated via ligation of the T cell receptor complex. 864 Aug 66

The serine/threonine protein kinase Raf-1 is a component of a conserved intracellular signaling cascade that controls responses to various extracellular stimuli. Transcription from several promoters, including the oncogene-responsive element in the polyomavirus enhancer, the c-fos promoter, as well as other AP-1- and Ets-dependent promoters, can be induced by Raf-1 kinase. Previously, we have shown that activated Raf-1 kinase transactivates the human immunodeficiency virus type 1 (HIV-1) long terminal repeat and have identified the NF-kappaB binding motif as a Raf-1-responsive element (RafRE). We now report that Raf-1 kinase-induced transactivation from the HIV RafRE involves the purine-rich-repeat-binding protein (GABP), which is composed of two distinct subunits (alpha and beta). GABP alpha is an Ets oncogene-related DNA-binding protein, and GABP beta contains four ankyrin-like repeats that have been shown to be essential in protein-protein interactions. In electrophoretic mobility shift assays using nuclear extracts from human Jurkat T cells, a protein-DNA complex which was supershifted with antiserum against GABP alpha and GABP beta was observed. Purified recombinant GABP alpha and beta interact with the HIV RafRE as judged from DNA binding assays. Cotransfection experiments with GABP alpha and beta and Raf-1 kinase demonstrate synergistic transactivation of the HIV-1 promoter. Point mutations in the HIV RafRE abolished the Raf-1 kinase as well as GABP alpha- and beta-induced transactivation. The observed Raf-1-GABP synergism presumably involves phosphorylation of GABP subunits, as treatment of cells with Raf-1 kinase activators serum and 12-O-tetradecanoylphorbol-13-acetate increases phosphorylation of GABP in vivo. However, GABP is not a target of Raf-1 kinase; instead, it is a substrate of mitogen-activated protein kinase (MAPK/ERK), since in vitro phosphorylation of GABP alpha and beta was achieved by the reconstituted protein kinase cascade but not with purified Raf-1 or MEK. These results suggest that Raf-1 kinase- induced activation of the HIV-1 promoter is mediated by the classical cytoplasmic cascade resulting in MAPK/ERK-mediated phosphorylation of GABP alpha and beta. Because the HIV RafRE corresponds to a region within the promoter which is essential for regulation of HIV-1 expression, the data indicate that in addition to NK-kappaB, GABP transcription factors are important for induced expression of HIV.
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PMID:Raf-1 kinase targets GA-binding protein in transcriptional regulation of the human immunodeficiency virus type 1 promoter. 864 52

A plethora of extracellular signals leads to the stimulation of Ras, which triggers intracellular protein kinase cascades, resulting in activation of transcription factors and thus in enhanced gene activity. In this report, it is demonstrated that the ETS transcription factor ER81, which appears to be localized within the cell nucleus by virtue of its DNA binding domain, is transcriptionally activated by oncogenic Ras. Since this activation was dependent on the presence of Raf-1 and ERK-1, ER81 is a target of the Ras/Raf/MEK/ERK signaling cascade. Consistently, activated ERK-1 is capable to phosphorylate ER81. However, the carboxy-terminal region of ER81, which contains no potential ERK phosphorylation sites, is also transcriptionally activated by ERK-1, suggesting that an ERK-stimulated protein kinase phosphorylates and thus stimulates ER81 activity. Two acidic stretches of amino acids, which are conserved in the related PEA3 and ERM proteins, are localized within the amino-and carboxy-terminal transactivation domains of ER81. In addition, an inhibitory domain may dampen the activation function of these two domains. In conclusion, ER81 is a target of Ras-dependent signaling cascades and may thus contribute to the nuclear response upon stimulation of cells and also to cellular transformation due to oncogenic Ras.
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PMID:Analysis of the ERK-stimulated ETS transcription factor ER81. 865 29

The chemotactic peptide f-Met-Leu-Phe (fMLP) stimulates leukocyte functions through binding and activation of a specific G-protein-coupled formyl peptide receptor (FPR). Recent studies have shown that stimulation of neutrophils with fMLP induces the activation of two members of the mitogen-activated protein kinase (MAP kinase) family, ERK1 and ERK2, through mechanisms that are not completely understood but may involve the phosphorylation of the adapter protein SHC by the Src-related kinase Lyn. In this study, transfected fibroblasts expressing the rabbit FPR were used to investigate further the role of Lyn and SHC phosphorylation in fMLP-stimulated MAP kinase activation. Stimulation of transfected cells with fMLP resulted in the time- and dose-dependent increase in tyrosine phosphorylation and activation of ERK1 and ERK2 and the activation of MEK, the MAP kinase/ERK kinase. The activation of both ERKs and MEK was inhibited by preincubation of the cells with pertussis toxin, indicating that activation was dependent upon a Gi/Go-like protein that couples to the receptor. Our data also show that, unlike neutrophils, FPR-transfected fibroblasts do not express the Src-related kinase Lyn. In the absence of Lyn, fMLP stimulation did not result in an increased tyrosine phosphorylation of the adapter protein SHC, whereas it was still able to induce MAP kinase activation. These data suggest that Lyn and SHC are not the only upstream signals for activation of the MAP kinase/ERK pathway by fMLP and demonstrate the potential application of the FPR-transfected cells for the delineation of additional signaling mechanisms stimulated by fMLP.
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PMID:Activation of the mitogen-activated protein kinase pathway by fMet-leu-Phe in the absence of Lyn and tyrosine phosphorylation of SHC in transfected cells. 866 60

Mitogen-activated protein (MAP) kinase cascades represent one of the major signal systems used by eukaryotic cells to transduce extracellular signals into cellular responses. Four MAP kinase subgroups have been identified in humans: ERK, JNK (SAPK), ERK5 (BMK), and p38. Here we characterize a new MAP kinase, p38beta. p38beta is a 372-amino acid protein most closely related to p38. It contains a TGY dual phosphorylation site, which is required for its kinase activity. Like p38, p38beta is activated by proinflammatory cytokines and environmental stress. A comparison of events associated with the activation of p38beta and p38 revealed differences, most notably in the preferred activation of p38beta by MAP kinase kinase 6 (MKK6), whereas p38 was activated nearly equally by MKK3, MKK4, and MKK6. Moreover, in vitro and in vivo experiments showed a strong substrate preference by p38beta for activating transcription factor 2 (ATF2). Enhancement of ATF2-dependent gene expression by p38beta was approximately20-fold greater than that of p38 and other MAP kinases tested. The data reported here suggest that while closely related, p38beta and p38 may be regulated by differing mechanisms and may exert their actions on separate downstream targets.
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PMID:Characterization of the structure and function of a new mitogen-activated protein kinase (p38beta). 866 24

The transcription factor, Nuclear Factor of Activated T cells (NFAT) is a major target for p21ras and calcium signalling pathways in the IL-2 gene and is induced by p21ras signals acting in synergy with calcium/calcineurin signals. One p21ras effector pathway involves the MAP kinase ERK-2, and we have examined its role in NFAT regulation. Expression of dominant negative MAPKK-1 prevents NFAT induction. Constitutively active MAPKK-1 fully activates ERK-2 and the transcription factor Elk-1, but does not substitute for activated p21ras and synergize with calcium/calcineurin signals to induce NFAT. Expression of dominant negative N17Rac also prevents TCR and p21ras activation of NFAT, but without interfering with the ERK-2 pathway. The transcriptional activity of the NFAT binding site is mediated by a complex comprising a member of the NFAT group and AP-1 family proteins. The induction of AP-1 by p21ras also requires Rac-1 function. Activated Rac-1 could mimic activated p21ras to induce AP-1 but not to induce NFAT. Moreover, the combination of activated MAPKK-1 and Rac-1 could not substitute for activated p21ras and synergize with calcium signals to induce NFAT. Thus, p21ras regulation of NFAT in T cells requires the activity of multiple effector pathways including those regulated by MAPKK-1/ERK-2 and Rac-1.
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PMID:Multiple p21ras effector pathways regulate nuclear factor of activated T cells. 867 Aug 97

SPRK (also called PTK-1 and MLK-3), a member of the mixed lineage kinase subfamily of (Ser/Thr) protein kinases, encodes an amino-terminal SH3 domain followed by a kinase catalytic domain, two leucine zippers interrupted by a short spacer, a Rac/Cdc42 binding domain, and a long carboxyl-terminal proline-rich region. We report herein that SPRK activates the stress-activated protein kinases (SAPKs) but not ERK-1 during transient expression in COS cells; the p38 kinase is activated modestly (1.3-2 fold) but consistently. SPRK also activates cotransfected SEK-1/MKK-4, a dual specificity kinase which phosphorylates and activates SAPK. Reciprocally, expression of mutant, inactive SEK-1 inhibits completely the basal and SPRK-activated SAPK activity. Immunoprecipitated recombinant SPRK is able to phosphorylate and activate recombinant SEK-1 in vitro to an extent comparable to that achieved by MEK kinase-1. These results identify SPRK as a candidate upstream activator of the stress-activated protein kinases, acting through the phosphorylation and activation of SEK-1.
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PMID:The mixed lineage kinase SPRK phosphorylates and activates the stress-activated protein kinase activator, SEK-1. 870 71


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