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Query: HUMANGGP:019459 (ERK2)
 3,473 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The activation of insulin-stimulated protein-serine/threonine kinases has been investigated in CHO cell lines transfected with cDNAs encoding either wild-type or mutant human insulin receptors. (1) Insulin treatment of CHO cells over-expressing wild-type insulin receptors resulted in the rapid and substantial (5-10-fold) activation of cytosolic protein kinases which phosphorylated myelin basic protein, Kemptide and two peptide substrates based on sites phosphorylated on ribosomal protein S6 in vivo. (2) Further fractionation of cytosolic extracts by MonoQ chromatography revealed two peaks of insulin-stimulated myelin basic protein kinase activity which were highly related to the previously described mitogen-activated protein (MAP) kinases ERK1 and ERK2. In addition, at least two major peaks of S6 kinase activity were resolved, which exhibited properties similar to the 70 kDa and 90 kDa S6 kinases described by others; the predominant effect of insulin was on the activity of the 90 kDa enzyme and was in excess of 10-fold. (3) MonoQ fractionation of extracts from parental CHO cells, or cells expressing kinase-deficient receptors, showed all insulin-stimulated peaks of activity to be almost completely absent. (4) Further studies demonstrated that substitution of tyrosine residues 1162 and 1163 (or 1162 alone) with phenylalanine led to a substantial reduction in the ability of insulin to stimulate these protein kinase activities when assayed in cytosolic extracts. In contrast, deletion of 69 amino acids from the C-terminus of the insulin receptor beta-subunit caused a leftward shift in the insulin dose-response curve of the MAP kinase activity, but apparently not in that of the 90 kDa S6 kinase activity.
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PMID:Characterization of insulin-stimulated protein serine/threonine kinases in CHO cells expressing human insulin receptors with point and deletion mutations. 132 27

Both MAP kinases and the protein kinase p74raf-1 are activated by many growth factors in a c-ras-dependent manner and by oncogenic p21ras. We were therefore interested in determining the relationship between MAP kinases and raf. The MAP kinase ERK2 is activated by expression of oncogenically activated raf, independently of cellular ras. Overexpressed p74raf-1 potentiates activation of ERK2 by EGF and TPA. MAP kinase kinase inactivated by phosphatase 2A treatment is phosphorylated and reactivated by incubation with p74raf-1 immunoprecipitated from phorbol ester-treated cells. We conclude that raf protein kinase is upstream of MAP kinases and is either a MAP kinase kinase kinase or a MAP kinase kinase kinase kinase.
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PMID:Activation of the MAP kinase pathway by the protein kinase raf. 133 Mar 21

Treatment of bovine chromaffin cells with nicotinic agonists, phorbol esters, and growth factors increases protein kinase activity toward microtubule-associated protein-2 and myelin basic protein (MBP) in vitro. To characterize the kinases that are activated by these agents, we separated chromaffin cell proteins by electrophoresis in sodium dodecyl sulfate-polyacrylamide gels into which MBP had been incorporated, allowed the proteins to renature, and then assayed MBP kinase activity by incubating the gels with [gamma-32P]ATP. Chromaffin cells contain a family of kinases that phosphorylate MBP in vitro. Two of these kinases, of M(r) 46,000 and 42,000 (PK46 and PK42), were activated by treatment of the cells with dimethylphenylpiperazinium (DMPP), phorbol 12,13-dibutyrate (PDBu), or insulin-like growth factor I (IGF-I). Activation of PK46 and PK42 by DMPP was dependent on extracellular Ca2+, whereas the effects of PDBu and IGF-I were Ca2+ independent. Down-regulation of protein kinase C by incubation of the cells with PDBu abolished the activation of PK46 and PK42 by DMPP, PDBu, and IGF-I. Staurosporine, a protein kinase C inhibitor, prevented the activation of PK46 and PK42 by DMPP and PDBu but did not block the activation of these kinases by IGF-I. Immunoblotting experiments with antiphosphotyrosine (anti-PTyr) antibodies demonstrated that agents that increased the kinase activities of PK46 and PK42 also increased the apparent PTyr content of M(r) 46,000 and 42,000 proteins. PK46 and PK42 comigrated with proteins that reacted with antibodies against extracellular signal-regulated kinases (ERKs). Thus, PK46 and PK42 appear to be the bovine homologues of ERK1 and ERK2.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Nicotinic agonists, phorbol esters, and growth factors activate two extracellular signal-regulated kinases, ERK1 and ERK2, in bovine chromaffin cells. 143 97

As detected by coimmunoprecipitation from PC12 cells, NGF induces rapid association between ERK1 (a growth factor-activated serine/threonine protein kinase) and gp140prototrk NGF receptors. In contrast, no such association is found with the closely related ERK2. Anti-trk immunocomplexes generated from NGF-treated cells also contain protein kinase activity that shares many properties with soluble ERK1. The association of both ERK1 protein and ERK-like kinase activity with gp140prototrk is maximal by 5 min of NGF treatment, persists for approximately 1 hr, and subsequently declines by 18 hr. Treatment with either basic fibroblast growth factor, epidermal growth factor, or orthovanadate also leads to association of ERK1 with gp140prototrk without tyrosine phosphorylation of the latter. The interaction between ERK1 and gp140prototrk may prove relevant to the NGF mechanism.
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PMID:NGF and other growth factors induce an association between ERK1 and the NGF receptor, gp140prototrk. 146 7

The small GTP-binding protein Ras appears to be required for transformation and differentiation induced by tyrosine kinases. The Ras requirement may be limited to a few tyrosine kinase-regulated signaling pathways or may be universal for all tyrosine kinase actions. Because both Ras and the microtubule-associated protein 2 kinases ERK1 and ERK2 have been implicated in events that lead to neurite outgrowth, we explored the possibility that Ras and ERKs may lie on the same signaling pathway. Utilizing PC-12 rat adrenal pheochromocytoma cell lines that contain a dominant inhibitory Ras mutant (S17N-Ras(H)), we found that Ras was required for stimulation of the ERK cascade by nerve growth factor but apparently not by the heterotrimeric G protein activator AlF4-. Within this cascade, Ras appears to be upstream of an ERK activator, raising the intriguing possibility that Ras may directly regulate a serine/threonine protein kinase.
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PMID:Evidence for a Ras-dependent extracellular signal-regulated protein kinase (ERK) cascade. 149 81

A protein kinase characterized by its ability to phosphorylate microtubule-associated protein-2 (MAP2) and myelin basic protein (MBP) is thought to play a pivotal role in the transduction of signals from many receptors in response to their ligands. A kinase with such activity, named extracellular signal-regulated kinase 1 (ERK1), is activated rapidly by numerous extracellular signals, requires phosphorylation on tyrosine to be fully active, and in vitro can activate a kinase (a ribosomal S6 protein kinase) that is downstream in phosphorylation cascades. From the protein sequence predicted by the rat ERK1 cDNA, peptides were synthesized and used to elicit antibodies. The antibodies recognize both ERK1; a closely related kinase, ERK2; and a third novel ERK-related protein. Using these antibodies we have determined that ERK1 and ERK2 are ubiquitously distributed in rat tissues. Both enzymes are expressed most highly in brain and spinal cord as are their mRNAs. The third ERK protein was found in spinal cord and in testes. The antibodies detect ERKs in cell lines from multiple species, including human, mouse, dog, chicken, and frog, in addition to rat, indicating that the kinases are conserved across species. ERK1 and ERK2 have been separated by chromatography on Mono Q. Stimulation by insulin increases the phosphorylation of both kinases on tyrosine residues, as assessed by immunoblotting with phosphotyrosine antibodies, and retards their elution from Mono Q. Each of these ERKs appears to account for a distinct peak of MBP kinase activity. The activity in each peak is diminished by incubation with either phosphatase 2a or CD45. Therefore, both enzymes have similar modes of regulation and appear to contribute to the growth factor-stimulated MAP2/MBP kinase activity measured in cell extracts.
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PMID:Identification of multiple extracellular signal-regulated kinases (ERKs) with antipeptide antibodies. 165 26

Microtubule-associated protein 2 kinase (MAP kinase), which exists in several forms, is a protein serine/threonine kinase that participates in a growth factor-activated protein kinase cascade in which it activates a ribosomal protein S6 kinase (pp90rsk) while being regulated itself by a cytoplasmic factor (MAP kinase activator). Experiments with recombinant MAP kinase, ERK2, purified from Escherichia coli in a nonactivated form revealed a self-catalyzed phosphate incorporation into both tyrosine and threonine residues. Another MAP kinase, ERK1, purified from insulin-stimulated cells also autophosphorylated on tyrosine and threonine residues. Autophosphorylation of ERK2 correlated with its autoactivation, although both autophosphorylation and autoactivation were slow compared to that occurring in the presence of MAP kinase activator. Therefore, we propose that autophosphorylation is probably involved in the MAP kinase activation process in vitro, but it may not be sufficient for full activation. The specificity toward tyrosine and threonine residues indicates that the MAP kinases ERK1 and ERK2 are members of a group of kinases with specificity for tyrosine as well as serine and threonine residues.
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PMID:Microtubule-associated protein 2 kinases, ERK1 and ERK2, undergo autophosphorylation on both tyrosine and threonine residues: implications for their mechanism of activation. 171 80

The Raf-1 proto-oncogene protein kinase can be phosphorylated and activated after stimulation of cells with insulin and a variety of other growth factors and mitogens. We recently presented evidence that insulin and certain other growth factors activated one or more Raf-1 kinase kinase activities (Lee, R.M., Rapp, U. R., and Blackshear, P.J. (1991) J. Biol. Chem. 266, 10351-10357). In the present study, four peaks of Raf-1 kinase kinase activity were identified after anion-exchange chromatography of cell lysates, and two of these were activated by insulin. Further chromatographic characterization of these two peaks of insulin-activated kinase activity indicated that they contained three apparently distinct kinase activities. Two of these activities comigrated with immunoreactive extracellular signal-regulated kinases (ERK) 1 and 2 (mitogen-activated protein kinase) through three different chromatographic separations. Both ERK1 and ERK2 phosphorylated Raf-1 with reasonably high affinity (Km for ERK1 = 90 nM; Km for ERK2 = 120 nM), and produced similar, complex phosphopeptide maps; both kinases also phosphorylated myelin basic protein. The third kinase activity also phosphorylated Raf-1 and myelin basic protein but did not comigrate exactly with either immunoreactive ERK1 or ERK2. We conclude that two and possibly three insulin-activated Raf-1 kinase kinases are members of the ERK family.
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PMID:Evidence that extracellular signal-regulated kinases are the insulin-activated Raf-1 kinase kinases. 173 Jun 37

The MAP kinase pathway impinging on ERK2 has been shown to be integrally associated with mitogenic signalling in many cell types. Previously, we and others have demonstrated that oncogenic forms of Raf-1 kinase, when expressed in fibroblasts, lead to the constitutive activation of ERK2, the de-regulation of c-fos expression and increased cell proliferation. Here we describe an exception to this scenario. In Rat6 cells, although both ERK1 and ERK2 are activated in response to mitogens that induce c-fos expression, such as Epidermal Growth Factor (EGF), lysophosphatidic acid (LPA) or serum, expression of v-Raf fails to induce c-fos expression and increase proliferation. However, ERK2 is activated by v-Raf expression. The co-transfection of an interfering mutant of ERK2 has no effect on the level of c-fos reporter expression in Rat6 cells whereas the analogous ERK1 mutant reduces its expression. Furthermore, the spontaneous focus formation observed in Rat6 cells is susceptible to the interfering mutant of ERK1 but resistant to that of ERK2. Thus, not only do mitogenic signals appear to by-pass both Raf-1 kinase and ERK2, the Raf-1-ERK2 pathway seems to be functionally compromised in Rat6 cells as its activation leads neither to c-fos expression nor to increased proliferation.
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PMID:Raf-1 kinase and ERK2 uncoupled from mitogenic signals in rat fibroblasts. 747 30

A consensus cyclic AMP response element (CRE) in the murine prostaglandin synthase-2 (PGS2) promoter is essential for pgs2 gene expression induced by pp60v-src, the v-src oncogene product. In this study, we investigate (i) the transcription factors active at the PGS2 "CRE site" in response to v-src activation and (ii) the signal transduction pathways by which pp60v-src activates these transcription factors. Transient transfection assays with pgs2 promoter/luciferase reporter chimeric genes suggest that c-Jun mediates v-src-induced pgs2 gene expression. Antibody supershift experiments demonstrate that c-Jun can participate in a complex with the pgs2 promoter CRE site. Moreover, in vitro immuno-complex assays demonstrate that pp60v-src expression strongly activates c-Jun N-terminal kinase (JNK1) enzyme activity. Serines 63 and 73, the sites of c-Jun phosphorylation by JNK, are essential for v-src-induced, pgs2 promoter-mediated luciferase expression. Cotransfection studies with plasmids expressing wild-type JNK, dominant-negative JNK, and dominant-negative MEKK-1 confirm that activation of the Ras/MEKK-1/JNK/c-Jun pathway is required for v-src-induced pgs2 gene expression. Overexpression of either wild-type ERK-1 or ERK-2 proteins also potentiate v-src-mediated luciferase expression driven by the pgs2 promoter, and expression of dominant-negative mutants of ERK-1, ERK-2, or Raf-1 attenuate this response. Thus, in response to v-src expression, a Ras/MEKK-1/JNK signal transduction pathway activating c-Jun and a Ras/Raf-1/ERK pathway converge to mediate pgs2 gene expression via the CRE site in the pgs2 promoter.
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PMID:v-src induces prostaglandin synthase 2 gene expression by activation of the c-Jun N-terminal kinase and the c-Jun transcription factor. 749 26


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