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:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

MEK, a dual specificity threonine/tyrosine kinase, has been postulated to be a convergent point for signaling from receptor protein tyrosine kinases (RTKs) and G-protein-coupled receptors. In contrast to yeast and mammalian cells where several MEKs have been isolated, only one Drosophila MEK (D-Mek) has been characterized to date. Previous studies have shown that D-Mek acts in the Torso RTK signaling pathway. To demonstrate that D-Mek also operates downstream of other RTKs, we generated a temperature-sensitive allele of D-mek (D-mekts) by site-directed mutagenesis based on the amino acid change of a yeast cdc2ts mutation. Using D-mekts, we show that in addition to its role in Torso signaling, D-Mek operates in the Sevenless and in the Drosophila epidermal growth factor RTK pathways. Because loss-of-function mutations in D-mek and the upstream receptors give rise to similar phenotypes, it suggests that D-mek is the only MEK activated by Drosophila RTKs. In addition, we demonstrate that different RTK pathways respond differently to alteration in D-Mek activity.
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PMID:A temperature-sensitive MEK mutation demonstrates the conservation of the signaling pathways activated by receptor tyrosine kinases. 795 87

Bacterial LPS is a potent macrophage activator. The early steps in LPS signal transduction involve the tyrosine phosphorylation and activation of a number of kinases of the src family, and inhibition of this pathway causes a severe impairment in the production of the cytokines TNF-alpha and IL-1 beta. We find that LPS-induced macrophages activation also involves the Raf-1 kinase, a key component in mitogenic signal transduction. Treatment of BAC-1.2F5 macrophages with LPS causes phosphorylation and activation of Raf-1. This is paralleled by the stimulation of MEK-1 and MAP-kinase activity and by the phosphorylation of the transcription factor Elk-1, a nuclear target of MAP-kinase. Activation of the Raf/MAP-kinase pathway was inhibited upon pretreatment of the cells with genistein, a tyrosine kinase inhibitor. Raf-1 must thus lie downstream of tyrosine kinase in LPS signal transduction. However, Raf-1 is not a direct substrate of a LPS-induced tyrosine kinase, because Raf-1 immunoisolated from LPS-induced cells contains only phosphoserine. This resembles the situation after CSF-1-stimulation of macrophages, in which Raf-1 clearly transduces a signal generated by the CSF-1 receptor kinase, but is phosphorylated exclusively in serine. Phosphopeptide maps of Raf-1 immunoprecipitated from LPS- or CSF-1-treated cells are indistinguishable, suggesting that these agents activate Raf-1 by similar mechanisms. Finally, v-raf-infected BAC-1.2F5 macrophages were found to constitutively express low levels of IL-1 beta and TNF-alpha. These data argue that Raf-1 functions downstream of tyrosine kinases in LPS-mediated macrophage activation and cytokine production.
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PMID:Lipopolysaccharide induces activation of the Raf-1/MAP kinase pathway. A putative role for Raf-1 in the induction of the IL-1 beta and the TNF-alpha genes. 798 71

Src homology/collagen (SHC) proteins are thought to participate in signaling through both receptor tyrosine kinases, such as the insulin receptor and the EGF (epidermal growth factor) receptor, and cytoplasmic tyrosine kinases, such as v-src and v-fps. Here we approached the insulin-induced and the insulin-like-growth-factor-I-induced (IGF-I-induced) phosphorylation of SHC proteins, and the possible role of these proteins in insulin and IGF-I signaling. First, we showed that SHC proteins are phosphorylated on tyrosine residues upon insulin and IGF-I treatment of fibroblasts transfected with a SHC cDNA construct. More important, ligand-activated insulin and IGF-I receptors phosphorylate SHC proteins in vitro, indicating that SHC proteins could be direct substrates for insulin and IGF-I receptors. Further, insulin or IGF-I treatment of SHC-transfected fibroblasts leads to immunoprecipitation of SHC proteins with insulin-receptor substrate 1 (IRS-1). We next looked at the possible effect of SHC proteins on biological responses in SHC-transfected fibroblasts. We found that the expression of exogenous SHC proteins results in an increased basal MEK (MAPK/ERK-activating kinase) activity. Further, neither the basal nor the insulin-induced or IGF-I-induced PtdIns-3-kinase activity were modified by expression of exogenous SHC proteins. These results illustrate that SHC proteins are implicated in the MAP (mitogen-activated protein)-kinase pathway, but not in that of PtdIns-3-kinase. Finally, we show that SHC-transfected cells, unlike control cells, are able to advance into the early phases of the cell cycle, and are more sensitive to the growth-promoting effect of insulin. In conclusion, SHC proteins are substrates for insulin and IGF-I receptors, and would appear to function as early post-receptor signaling components.
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PMID:Involvement of Src-homology/collagen (SHC) proteins in signaling through the insulin receptor and the insulin-like-growth-factor-I-receptor. 803 92

Chemoattractants bind to seven transmembrane-spanning, G-protein-linked receptors on polymorphonuclear leukocytes (neutrophils) and induce a variety of functional responses, including activation of microtubule-associated protein (MAP) kinase. Although the pathways by which MAP kinases are activated in neutrophils are unknown, we hypothesized that activation of the Ras/Raf pathway leading to activation of MAP/ERK kinase (MEK) would be induced by the chemoattractant f-met-leu-phe. Human neutrophils exposed to 10 nM FMLP for 30 s exhibited an MAP kinase kinase activity coeluting with MEK-1. Immunoprecipitation of Raf-1 kinase after stimulation with FMLP revealed an activity that phosphorylated MEK, was detectable at 30 s, and peaked at 2-3 min. Immunoprecipitation of Ras from both intact neutrophils labeled with [32P]orthophosphate and electropermeabilized neutrophils incubated with [32P]GTP was used to determine that FMLP treatment was associated with activation of Ras. Activation of both Ras and Raf was inhibited by treatment of neutrophils with pertussis toxin, indicating predominant linkage to the Gi2 protein. Although phorbol esters activated Raf, activation induced by FMLP appeared independent of protein kinase C, further suggesting that Gi2 was linked to Ras and Raf independent of phospholipase C and protein kinase C. Dibutyryl cAMP, which inhibits many neutrophil functional responses, blocked the activation of Raf by FMLP, suggesting that interruption of the Raf/MAP kinase pathway influences neutrophil responses to chemoattractants. These data suggest that Gi2-mediated receptor regulation of the Ras/Raf/MAP kinase pathway is a primary response to chemoattractants.
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PMID:FMLP activates Ras and Raf in human neutrophils. Potential role in activation of MAP kinase. 804 Feb 99

The simian virus 40 small tumor antigen (small t) specifically interacts with protein phosphatase type 2A (PP2A) in vivo and alters its catalytic activity in vitro. Among the substrates for PP2A in vitro are the activated forms of MEK and ERK kinases. Dephosphorylation of the activating phosphorylation sites on MEK and ERKs by PP2A in vitro results in a decrease in their respective kinase activities. Recently, it has been shown that overexpression of small t in CV-1 cells results in an inhibition of PP2A activity toward MEK and ERK2 and a constitutive upregulation of MEK and ERK2 activity. Previously, we have observed that overexpression of either ERK1, MEK1, or a constitutively active truncated form of c-Raf-1 (BXB) is insufficient to activate AP-1 in REF52 fibroblasts. We therefore examined whether overexpression of small t either alone or in conjunction with ERK1, MEK1, or BXB could activate AP-1. We found that coexpression of small t and either ERK1, MEK1, or BXB resulted in an increase in AP-1 activity, whereas expression of either small t or any of the kinases alone did not have any effect. Similarly, coexpression of small t and ERK1 activated serum response element-regulated promoters. Coexpression of kinase-deficient mutants of ERK1 and ERK2 inhibited the activation of AP-1 caused by expression of small t and either MEK1 or BXB. Coexpression of an interfering MEK, which inhibited AP-1 activation by small t and BXB, did not inhibit the activation of AP-1 caused by small t and ERK1. In contrast to REF52 cells, we observed that overexpression of either small or ERK1 alone in CV-1 cells was sufficient to stimulate AP-1 activity and that this stimulation was not enhanced by expression of small t and ERK1 together. These results show that the effects of small t on immediate-early gene expression depend on the cell type examined and suggest that the mitogen-activated protein kinase activation pathway is distinctly regulated in different cell types.
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PMID:Simian virus 40 small t antigen cooperates with mitogen-activated kinases to stimulate AP-1 activity. 806 56

The protein kinase cascade Raf-MAPKK/MEK-MAPK/ERK connects protein tyrosine kinase receptors in the membrane with control of transcription factor activity in the nucleus. We have examined whether Raf is obligatory for activation of this cascade and whether this signaling pathway is relevant to transformation. By use of transient assays with epitope-tagged ERK-1 cDNA and a dominant inhibitory mutant of Raf-1 we found that serum and 12-O-tetradecanoylphorbol-13-acetate as well as representatives of three classes of oncogenes (protein tyrosine kinases abl/src, Ras, and protein serine/threonine kinases mos/cot) were all Raf-dependent for stimulation of MAPK. All of the MAPK stimulating oncogenes were also activators of Raf kinase as judged by shift induction. It thus appears that there is little or no redundancy in pathways used by growth regulators for activation of MAPK/ERK. Furthermore, the ability to stimulate MAPK/ERK appears to be critical for transformation by oncogenic Raf-1 and ERK-1 and -2 synergized with v-raf in a focus induction assay on NIH3T3 cells and kinase dead mutants of ERK-2 were inhibitory. Raf/ERK synergism was also observed in transcriptional transactivation of the oncogene-response element in the polyoma enhancer. We conclude that this Raf signaling pathway, which connects to many upstream activators and downstream effectors, is essential for transformation by most oncogenes.
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PMID:Mitogen-activated protein kinase/extracellular signal-regulated protein kinase activation by oncogenes, serum, and 12-O-tetradecanoylphorbol-13-acetate requires Raf and is necessary for transformation. 812 67

Stimulation of T cells with antibodies directed towards the T cell receptor complex results in the activation of mitogen-associated protein kinase (MAPK). Two pathways have been described in other cell types that can lead to MAPK activation. One of these pathways involves the activation of Ras, leading to the activation of Raf-1, and the subsequent activation of MEK (MAPK or ERK kinase). The contribution of this pathway in T cells for anti-CD3 or phorbol myristate acetate (PMA)-mediated MAPK activation was examined. We detected the kinase activities of Raf-1 and MEK towards their substrates (MEK for Raf-1 and MAPK for MEK) in this pathway leading to the activation of MAPK. Stimulation of the T cells with either anti-CD3 antibody or PMA resulted in a rapid activation of both Ras and Raf-1. MEK activity towards kinase-active or -inactive recombinant MAPK also increased upon stimulation. In addition, both MAPK and p90rsk were activated in these cells. We suggest that activation of MAPK and the subsequent activation of ribosomal S6 kinase (p90rsk) occurs by the Ras/Raf-1/MEK cascade in T lymphocytes stimulated by ligation of the T cell receptor complex.
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PMID:Ligation of the T cell receptor complex results in activation of the Ras/Raf-1/MEK/MAPK cascade in human T lymphocytes. 818 45

The mitogen-activated protein (MAP) kinases are serine-threonine protein kinases that are activated by tyrosine and threonine phosphorylation by the dual specificity protein kinase MEK (MAP kinase/ERK kinase). The present report describes the purification to near homogeneity and characterization of a protein tyrosine phosphatase from Xenopus laevis eggs that dephosphorylates MAP kinase phosphorylated by MEK. Bacterially expressed Xenopus MAP kinase phosphorylated by purified Xenopus MEK was used as substrate throughout the purification. The purification procedure included anion-exchange, cation-exchange, gel filtration, heparin-Sepharose, and chromatography on a column of thiophosphorylated MAP kinase-Sepharose, resulting in a > 3000-fold purification. Upon analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, a protein of 47 kDa correlated with activity. The phosphatase showed absolute specificity toward phosphotyrosine and no activity toward phosphothreonyl-phosphoseryl residues of MAP kinase. The pH optimum of the enzyme was 7.0 with a Km of 9.0 microM for phosphorylated MAP kinase. The phosphatase was inhibited by ammonium molybdate (IC50, 2 microM), vanadate (IC50, 250 microM), millimolar concentrations of MnCl2, ZnCl2 and p-nitrophenylphosphate but not by okadaic acid or microcystin. This tyrosine phosphatase may be involved in deactivating MAP kinase in vivo.
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PMID:Purification and characterization of a mitogen-activated protein kinase tyrosine phosphatase from Xenopus eggs. 822 71

We have used the polymerase chain reaction and Northern blotting to identify protein tyrosine kinases that may play an important role in the process of melanoma initiation and progression. Degenerate primers from the conserved catalytic domain of tyrosine kinase genes were used to amplify and clone partial cDNA sequences from a human melanoma cell line (DX3-LT5.1) and normal human melanocytes. When the melanoma reaction products were sequenced, 13 distinct clones were found, of which one is novel to date and has provisionally been named MEK (for melanocytic kinase). Of the remaining 12 known kinases, only two, ERB-B2 and IGF1-R, have previously been reported in pigment cells. Reaction products from melanocytes included only eight of these 13 sequences. To test for quantitative differences in tyrosine kinase expression between normal and malignant cells, a panel of eight melanoma lines and normal melanocytes was analyzed by Northern blotting. Two tyrosine kinases (JTK-14/TIE and TYRO-9) were detected in some melanomas but were not found in normal melanocytes, whereas others, including MEK, appeared to be overexpressed in some malignant lines. A minority of kinases showed either no change or a reduction in the level of mRNA. Expression of tyrosine kinases varied independently, and individual lines contained various combinations of these enzymes. Our findings are consistent with an increased overall expression of these putative growth factor receptors during melanoma development.
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PMID:Novel and known protein tyrosine kinases and their abnormal expression in human melanoma. 822 28

Interaction with SV40 small tumor antigen (small t) compromised the ability of multimeric protein phosphatase 2A to inactivate the mitogen-activated protein kinase ERK1 and the mitogen-activated protein kinase kinase MEK1. Transient expression of small t in CV-1 cells activated MEK and ERK but did not affect Raf activity. Small t stimulated the growth of quiescent CV-1 cells almost as effectively as did serum. Coexpression of kinase-deficient ERK2 blocked most, but not all, of the proliferation caused by small t. Activation of the mitogen-activated protein kinase pathway and stimulation of cell growth were dependent on the interaction of small t with protein phosphatase 2A. These findings indicate that SV40 small t is capable of inducing cell growth through blockade of protein phosphatase and deregulation of the mitogen-activated protein kinase cascade.
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PMID:The interaction of SV40 small tumor antigen with protein phosphatase 2A stimulates the map kinase pathway and induces cell proliferation. 825 25


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