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:2.7.11.24 (mitogen-activated protein kinase)
95,810 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We demonstrate that members of the erk-encoded family of mitogen-activated protein (MAP) kinases (pp44/42mapk/erk) and members of the rsk-encoded protein kinases (RSKs or pp90rsk) are present in the cytoplasm and nucleus of HeLa cells. Addition of growth factors to serum-deprived cells results in increased tyrosine and threonine phosphorylation and in the activation of cytosolic and nuclear MAP kinases. Activated MAP kinases then phosphorylate (serine/threonine) and activate RSKs. Concurrently, a fraction of the activated MAP kinases and RSKs enter the nucleus. In addition, a distinct growth-regulated RSK-kinase activity (an enzyme[s] that phosphorylates recombinant RSK in vitro and that may be another member of the erk-encoded family of MAP kinases) was found associated with a postnuclear membrane fraction. Regulation of nuclear MAP kinase and RSK activities by growth factors and phorbol ester is coordinate with immediate-early gene expression. Indeed, in vitro, MAP kinase and/or RSK phosphorylates histone H3 and the recombinant c-Fos and c-Jun polypeptides, transcription factors phosphorylated in a variety of cells in response to growth stimuli. These in vitro studies raise the possibility that the MAP kinase/RSK signal transduction pathway represents a protein-Tyr/Ser/Thr phosphorylation cascade with the spatial distribution and temporal regulation that can account for the rapid transmission of growth-regulating information from the membrane, through the cytoplasm, and to the nucleus.
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PMID:Nuclear localization and regulation of erk- and rsk-encoded protein kinases. 154 23

This study characterizes the insulin-activated serine/threonine protein kinases in H4 hepatoma cells active on a 37-residue synthetic peptide (called the SKAIPS peptide) corresponding to a putative autoinhibitory domain in the carboxyl-terminal tail of the p70 S6 kinase as well as on recombinant p70 S6 kinase. Three peaks of insulin-stimulated protein kinase active on both these substrates are identified as two (possibly three) isoforms of the 40-45-kDa erk/microtubule-associated protein (MAP)-2 kinase family and a 150-kDa form of cdc2. Although distinguishable in their substrate specificity, these protein kinases together with the p54 MAP-2 kinase share a major common specificity determinant reflected in the SKAIPS peptide: the requirement for a proline residue immediately carboxyl-terminal to the site of Ser/Thr phosphorylation. In addition, however, at least one peak of insulin-stimulated protein kinase active on recombinant p70, but not on the SKAIPS peptide, is present although not yet identified. MFP/cdc2 phosphorylates both rat liver p70 S6 kinase and recombinant p70 S6 kinase exclusively at a set of Ser/Thr residues within the putative autoinhibitory (SKAIPS peptide) domain. erk/MAP kinase does not phosphorylate rat liver p70 S6 kinase, but readily phosphorylates recombinant p70 S6 kinase at sites both within and in addition to those encompassed by the SKAIPS peptide sequences. Although the tryptic 32P-peptides bearing the cdc2 and erk/MAP kinase phosphorylation sites co-migrate with a subset of the sites phosphorylated in situ in insulin-stimulated cells, phosphorylation of the p70 S6 kinase by these proline-directed protein kinases in vitro does not reproducibly activate p70 S6 kinase activity. Thus, one or more erk/MAP kinases and cdc2 are likely to participate in the insulin-induced phosphorylation of the p70 S6 kinase. In addition to these kinases, however, phosphorylation of the p70 S6 kinase by other as yet unidentified protein kinases is necessary to recapitulate the multisite phosphorylation required for activation of the p70 S6 kinase.
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PMID:An array of insulin-activated, proline-directed serine/threonine protein kinases phosphorylate the p70 S6 kinase. 173 88

Treatment of Swiss mouse 3T3 fibroblasts with certain cyclic nucleotide phosphodiesterase inhibitors (theophylline, SQ 20,006, and MY-5445) prevents the activation of the M(r) 70,000 S6 kinase (p70S6k) induced by a variety of external stimuli. Concentrations giving half-maximal inhibition were 800, 50, and 25 microM, respectively. Western blot analysis and immunocomplex kinase assays showed that these compounds inhibit the phosphorylation and activation of p70S6k without affecting the erk-encoded mitogen-activated protein (MAP) kinases or the rsk-encoded S6 kinase (p90rsk). A distinct collection of cAMP and cGMP agonists and analogues did not suppress p70S6k activation, indicating that 1) high intracellular cyclic nucleotide concentrations do not antagonize the p70S6k pathway and 2) phosphodiesterase inhibitors block p70S6k activation by a mechanism that is independent of cAMP or cGMP production. The effect of theophylline and SQ 20,006, but not MY-5445, on p70S6k signaling may be due in part to the inhibition of a phosphatidylinositol 3-kinase that acts upstream of p70S6k. Finally, in contrast to many other cell types, cAMP and cGMP were also found to have no inhibitory effect on the MAP kinase/p90rsk signaling pathway in Swiss 3T3 fibroblasts.
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PMID:Activation of p70 S6 kinase and erk-encoded mitogen-activated protein kinases is resistant to high cyclic nucleotide levels in Swiss 3T3 fibroblasts. 759 86

The metabolic and mitogenic actions of insulin have been proposed to be mediated by cellular serine/threonine kinases such as the ribosomal protein S6 kinases pp70-S6 (pp70-S6 kinase) and pp90rsk and the erk-encoded mitogen-activated protein kinases (pp42mapk and pp44mapk). Rapamycin completely blocked activation of pp70-S6 kinase by insulin in 3T3-L1 adipocytes, but did not inhibit insulin-stimulated glucose transport, translocation of GLUT4 to the cell surface, or activation of pp90rsk or pp44mapk by insulin. Concordant with the inhibition of kinase activity, rapamycin prevented the insulin-induced decrease in mobility of pp70-S6 kinase visualized by SDS-polyacrylamide gel electrophoresis, reflecting a reduction in the hormone-stimulated phosphorylation of the enzyme. The structurally related macrolide, FK506, had no effect on pp70-S6 kinase or hexose uptake. These data demonstrate that rapamycin blocks insulin activation of pp70-S6 kinase in 3T3-L1 adipocytes and that pp70-S6 kinase is not required in the signaling pathway leading to insulin-stimulated glucose transport.
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PMID:Dissociation of pp70 ribosomal protein S6 kinase from insulin-stimulated glucose transport in 3T3-L1 adipocytes. 767 6

Phorbol ester tumor promoters (TPA) activate the endogenous erk/MAP kinases and Rsk S6 kinases but not the p70S6 kinase in COS cells. DNA sequences encoding the rat Rsk-1 S6 kinase (homologous to Xenopus rsk alpha), modified by insertion of a peptide epitope at the polypeptide aminoterminus, were expressed transiently in COS cells. TPA stimulates the 40S and peptide kinase activity of the recombinant epitope-tagged Rsk-1, as well as the extent of Rsk-1 autophosphorylation in vitro (32P-Ser >> 32P-Thr). Indications that the conformation of the recombinant Rsk-1 polypeptide is substantially changed after activation by TPA in situ include a retarded mobility of the Rsk-1 polypeptide on SDS-PAGE and the appearance of new 32P-peptides during autophosphorylation in vitro. All these features of the TPA-activated Rsk-1 S6 kinase are abolished by dephosphorylation of the kinase in vitro with Ser/Thr phosphatase-2A. TPA increases 32P incorporation into recombinant Rsk-1 by 2-3-fold (32P-Ser >> 32P-Thr). Peptide mapping exhibits a single major 32P-peptide in Rsk-1 isolated from unstimulated cells and 10-12 additional 32P peptides after TPA treatment in situ. Phosphorylation of basal or phosphatase-2A-treated recombinant Rsk-1 in vitro with erk2/MAP kinase increases Rsk-1 40S kinase, peptide kinase, and autophosphorylating activity, retards migration of Rsk-1 polypeptides on SDS-PAGE, and generates new sites of Rsk-1 autophosphorylation in vitro. By contrast, TPA-activated Rsk-1 is not altered in these properties by autophosphorylation in vitro. By contrast, TPA-activated Rsk-1 is not altered in these properties by phosphorylation in vitro with erk2/MAP kinase. Activation of Rsk-1 in situ with TPA diminishes by over 90% the extent of Rsk-1 phosphorylation achieved in vitro by erk2/MAP kinase, as compared to the parallel phosphorylation of a phosphatase-2A-treated Rsk-1; basal Rsk-1 is intermediate. Peptide maps of phosphatase-2A-treated Rsk-1 after phosphorylation in vitro with erk2/MAP kinase exhibit 32P-peptides that comigrate with nearly all of the 32P-peptides present in TPA-activated-32P Rsk-1 labeled in situ, plus several 32P-peptides characteristic of Rsk-1 autophosphorylation in vitro.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Regulation of an epitope-tagged recombinant Rsk-1 S6 kinase by phorbol ester and erk/MAP kinase. 768 67

Lysophosphatidic acid (LPA) was shown to be a powerful inhibitor of gap-junctional communication between cultured rat liver WB cells, as determined by the transfer of Lucifer Yellow, with 50% inhibition obtained at about 0.3 microM LPA. Inhibition of communication was rapid (5 min) and was maintained for at least 80 min. After incubation for 3 h with LPA, communication competence was partially restored and dye transfer was refractory to further addition of LPA. Communication in LPA-refractory cells retained sensitivity to inhibition by phorbol ester and by epidermal growth factor (EGF). LPA-induced inhibition was associated with phosphorylation of connexin-43 protein, as detected by slower migration of the protein detected on Western blots, which could be eliminated by incubation of samples with alkaline phosphatase. A close correspondence was observed between the time- and dose-dependency of LPA effects on communication and the induction of mitogen-activated protein kinase (MAP kinase). Activation of both the 42 kDa and 44 kDa subspecies were confirmed by mobility shifts on Western blots using an anti-(MAP kinase R1) (erk 1-III) antibody and by fractionation on Mono Q columns. Cells pretreated with phorbol ester for 24 h were insensitive to phorbol ester inhibition of communication or activation of MAP kinase, but retained their sensitivity to LPA. The results indicate that LPA initiates the activation of protein kinase cascades in WB cells that are probably independent of protein kinase C and identifies connexin-43 as one substrate for the activated kinases.
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PMID:Lysophosphatidic acid inhibits gap-junctional communication and stimulates phosphorylation of connexin-43 in WB cells: possible involvement of the mitogen-activated protein kinase cascade. 798 Apr 7

Insulin exerts diverse effects on mitogenesis, metabolism, gene expression, and protein synthesis depending on the target cell type. A variety of extracellular serine/threonine kinases, including the ribosomal protein S6 kinases pp70-ribosomal S6 kinase (pp70-S6K) and pp90-ribosomal S6 kinase (pp90rsk) and the erk-encoded mitogen-activated protein (MAP) kinases pp44mapk/ERK-1 and pp42mapk/ERK-2, have been postulated as mediators of insulin action. In this study, we have investigated the role of the MAP kinase/pp90rsk signaling pathway in insulin-stimulated glucose transport in 3T3-L1 adipocytes. Differentiation of 3T3-L1 fibroblasts into adipocyte-like cells was accompanied by a marked increase in the capacity of insulin to activate pp90rsk and pp44mapk. Whereas the maximal insulin-stimulated pp90rsk and pp44mapk activities were only approximately 30% of the serum-stimulated activities in preadipocytes, the insulin-stimulated kinase activities in adipocytes were equal to or greater than the serum-stimulated activities. The increase in hormone receptor number accompanying differentiation accounted for the greater sensitivity, as overexpression of human insulin receptors in NIH-3T3 cells also conferred insulin-stimulatable kinase activity. In 3T3-L1 adipocytes, the stimulation of pp90rsk and pp44mapk activities was sufficiently rapid and hormone sensitive to convey a signal for increased hexose uptake. However, epidermal growth factor and fetal bovine serum were equipotent with insulin in stimulating pp90rsk and pp44mapk activities in adipocytes, but were without effect on hexose uptake. These data indicate that activation of these enzymes is not sufficient for the acute stimulation of glucose transport.
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PMID:Characterization of the mitogen-activated protein kinase/90-kilodalton ribosomal protein S6 kinase signaling pathway in 3T3-L1 adipocytes and its role in insulin-stimulated glucose transport. 829 68

The growth-promoting effects of endothelin-1 (ET-1) were examined in adult heart cells. The activity of mitogen-activated protein kinases (MAPKs) was measured in cytosolic extracts of isolated adult feline cardiac myocytes incubated with and without ET-1. Kinase activity was assessed by phosphorylation of the exogenous substrate, myelin basic protein. ET-1 stimulated the activity of MAPK up to 4-fold, with peak activation occurring between five and ten minutes after addition of ET-1. Polyclonal antisera raised against a 14-amino acid sequence of the erk-2 gene product, a MAPK isoform, identified two major bands in cytosolic extracts of the cardiac myocytes. Partial purification of kinase activities using Mono Q anion-exchange chromatography demonstrated two major peaks of myelin basic protein kinase activity. Subsequent immunoblots of the eluted fractions demonstrated that the immunoreactive bands observed in the cytosolic extracts eluted in those fractions possessing kinase activity. Overnight pretreatment of the cardiac myocytes with 100 ng/ml pertussis toxin inhibited the ET-1 stimulated increase in MAPK activity by 50 - 70%, but did not alter stimulation by 100 nM phorbol 12-myristate 13-acetate (PMA). These data suggest that stimulation of MAPK by ET-1 may be mediated by more than one pathway. MAPK has been shown to be activated in the intracellular transmission of growth factor signals. Indicative of a growth effect in this adult heart cell model, myocytes exposed to increasing concentrations of ET-1 demonstrated a dose dependent increase in [3H]-phenylalanine incorporation into cellular protein. This response was blocked by staurosporine and partially inhibited by pretreatment with pertussis toxin, again suggesting the possible involvement of multiple early signals. These data from isolated adult cardiac myocytes further support the hypothesis that ET-1 has a role in the regulation of cardiac growth.
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PMID:Effects of endothelin on mitogen-activated protein kinase activity and protein synthesis in isolated adult feline cardiac myocytes. 863 15

Human vascular endothelial cells (HUVECs), which do not display the lipopolysaccharide (LPS) receptor CD14, were examined for protein tyrosine phosphorylation after LPS stimulation in the presence and absence of soluble CD14 (sCD14). By phosphotyrosine Western blotting and immunocomplex kinase assays we show that LPS was capable of inducing in these cells rapid protein tyrosine phosphorylation and kinase activation of two members of the mitogen-activated protein kinase (MAPK) family erk-1 and the newly discovered p38, requiring the presence of sCD14. LPS-induced tyrosine phosphorylation of MAPK was associated with increased transcript- and surface protein expression of intracellular adhesion molecule-1 by HUVECs. MAPK phosphorylation and activation was induced by LPS in concentrations as little as 30 ng/mL and as early as 15 minutes after stimulation. Furthermore, tyrosine kinase inhibitors such as Genistein partially inhibited this effect. These results show that LPS triggers similar signaling events in both CD14+ myelo-monocytic cells and cells lacking the putative LPS-receptor CD14, suggesting the presence of a common, yet unidentified element in LPS-signaling in both cell types.
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PMID:Lipopolysaccharide induces the rapid tyrosine phosphorylation of the mitogen-activated protein kinases erk-1 and p38 in cultured human vascular endothelial cells requiring the presence of soluble CD14. 863 98

When HD3 colon carcinoma cells differentiate to fluid-transporting, enterocytic-like cells, they down-regulate their protein kinase C (PKC) beta levels 5-10-fold and lose two responses to basic fibroblast growth factor (FGF): proliferation and the ability to activate p57 mitogen-activated protein (MAP) kinase. HD3 cells were transfected with expression plasmids for the splice variants PKC-beta 1 and PKC-beta 2 and the empty vector for a control. Each of two PKC-beta 1 and each of two PKC-beta 2 transfectant clones exhibited elevated levels of Ca(2+)-and phosphatidylserine-dependent PKC activity. Both PKC-beta 1 transfectant clones had elevated levels of PKC-beta 1 protein compared with the PKC-beta 2 transfectants or controls, whereas both PKC-beta 2 transfectant clones had elevated levels of PKC-beta 2 protein compared with PKC-beta 1 transfectants. Control transfectants had no detectable PKC-beta 2 protein. Similar levels of PKC-alpha were found in all lines. Each PKC-beta transfectant was less differentiated than the parental line and had regained proliferative response to basic FGF. Increased growth rates in athymic mice were seen for PKC-beta 2 and PKC-beta 1 transfectant cells. Immunocytochemistry of the sectioned tumors showed enhanced protein levels of PKC-beta 2 and PKC-beta 1, correlating increased levels of these isonzymes with increased growth. Increased myelin-basic protein (MBP) kinase activities of M(r) 44,000, 57,000, 63,000, 110,000, and 130,000 by in-gel kinase assay characterized each PKC-beta transfectant. Both Western blotting and immunoprecipitation studies from 35S-prelabeled cells with a pan-erk antibody showed no increase in protein abundance of MAP kinases of M(r) 44,000, 57,000, and 63,000, suggesting that elevated PKC-beta levels led to activation of the smaller three MAP and MBP kinases. Activation of p57 MAP kinase in each PKC-beta transfectant was demonstrated by immunoprecipitation with an antiphosphotyrosine monoclonal antibody and then by assay of the immunoprecipitates by in-gel kinase assay on MBP. p57 MAP kinase was distinguished from the M(r) 54,000 stress-activated protein kinases, which migrated more rapidly on SDS gels and could be detected by in-gel kinase assay on MBP only after cellular stress. Thus, expression of elevated levels of PKC-beta 1 and PKC-beta 2 in differentiated HD3 colon carcinoma cells blocked their differentiation, enabled them to proliferate in response to basic FGF like undifferentiated cells, increased their growth rate in athymic mice, and activated several MBP kinases, among them, p57 MAP kinase.
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PMID:Protein kinase C beta 1 and protein kinase C beta 2 activate p57 mitogen-activated protein kinase and block differentiation in colon carcinoma cells. 873 68


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