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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)

Mitogen-activated protein kinases (MAP kinases) are activated by dual tyrosine and threonine phosphorylations in response to various stimuli, including phorbol esters. To define the mechanism of activation, recombinant wild-type 42-kDa MAP kinase (p42mapk) and a kinase-defective mutant of p42mapk (K52R) were used to assay both activator activity for p42mapk and kinase activity toward K52R in stimulated EL4.I12 mouse thymoma cells. Phorbol 12,13-dibutyrate (10 min, 650 nM) stimulated a single peak of MAP kinase activator that was coeluted from Mono Q at pH 7.5 and 8.9 with K52R kinase activity. Both activities were inactivated by the serine/threonine-specific phosphatase 2A but not by the tyrosine-specific phosphatase CD45. Phosphorylation of K52R occurred specifically on Thr-183 and Tyr-185, as determined by tryptic phosphopeptide mapping in comparison with synthetic marker phosphopeptides. These findings indicate that phorbol ester-stimulated MAP kinase kinase can activate p42mapk by threonine and tyrosine phosphorylations, and that p42mapk thus does not require an autophosphorylation reaction.
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PMID:The phorbol ester-dependent activator of the mitogen-activated protein kinase p42mapk is a kinase with specificity for the threonine and tyrosine regulatory sites. 131 55

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

Ligation of CD3/TCR on T-cells induces transient activation of lymphoid MAP-2 kinase (MAP-2K), a 43 kDa serine kinase which itself is a substrate of an unidentified tyrosine kinase (pp43). The reversibility of the MAP-2K response agrees with removal of tyrosine phosphates from pp43. Since both activity as well as tyrosine phosphorylation of MAP-2K could be prolonged by Na3VO4, a phosphotyrosine phosphatase inhibitor, we studied the effect of the common CD45 isoform, which is a member of the CD45 phosphatase family, on MAP-2K activity in vivo and in vitro. We demonstrate the ability of purified CD45 phosphatase to remove tyrosine phosphates from partially purified lymphoid MAP-2K. Utilizing the approach of heterologous receptor aggregation, we also showed that CD45 could inhibit the induction of MAP-2K activity in intact Jurkat cells during CD3 or CD3 + CD4 stimulation. We therefore suggest that this phosphatase may control the activity of lymphoid MAP-2K in vivo.
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PMID:Evidence for involvement of glycoprotein-CD45 phosphatase in reversing glycoprotein-CD3-induced microtubule-associated protein-2 kinase activity in Jurkat T-cells. 171 Aug 91

The most prominent tyrosyl-phosphorylated protein in maturing sea star oocytes was identified as the 44 kDa myelin basic protein (MBP) kinase p44mpk. Immunoblotting studies with anti-phosphotyrosine PY-20 antibody and phosphoamino acid analysis of in vivo [32P]phosphate-labelled p44mpk showed that the tyrosyl phosphorylation of the kinase correlated with a greater than 10-fold stimulation of its MBP phosphotransferase activity. The activation of p44mpk was reversed almost completely by purified preparations of the protein-tyrosyl phosphatases CD45 and 1B. Purified p44mpk has previously been shown to undergo autophosphorylation in vitro on seryl residues and this was associated with further enhancement of its MBP phosphorylating activity (Sanghera et al. (1991) J. Biol. Chem. 266, 6700-6707). p44mpk also underwent seryl phosphorylation during oocyte maturation, and the protein-seryl/threonyl phosphatase 2A reversed partially the maturation-associated stimulation of its MBP kinase activity. The properties of p44mpk resemble the murine 42 kDa mitogen-activated protein kinase (p42mapk). While p44mpk may feature the phosphorylatable tyrosyl residue that is critical for activation in p42mapk, it lacks the upstream threonyl phosphorylation site that is also required for p42mapk activity (Payne et al. (1991) EMBO J: 10, 885-892). These findings indicate partial differences in the regulatory mechanisms that govern the activities of these isozymes.
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PMID:Tyrosyl phosphorylation and activation of the myelin basic protein kinase p44mpk during sea star oocyte maturation. 171 36

MAP kinase (relative molecular mass, 42,000), a low abundance serine--threonine protein kinase, is transiently activated in many cell types by a variety of mitogens, including insulin, epidermal growth factor, and phorbol esters. In vitro, MAP kinase will phosphorylate and reactivate S6 kinase II previously inactivated by phosphatase treatment. Because many of the stimuli that activate MAP kinase are also stimulators of cell proliferation, and regulation of the cell cycle seems to involve a network of protein kinases, MAP kinase could be important in the transmission of stimuli eventually leading to the progression from G0 to G1 in the cell cycle. Activated MAP kinase contains both phosphotyrosine and phosphothreonine. We report here that MAP kinase can be deactivated completely by treatment with either phosphatase 2A, a protein phosphatase specific for phosphoserine and phosphothreonine, or CD45, a phosphotyrosine-specific protein phosphatase. We demonstrate that MAP kinase is only active when both tyrosyl and threonyl residues are phosphorylated and suggest therefore that the enzyme functions in vivo to integrate signals from two distinct transduction pathways.
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PMID:Requirement for integration of signals from two distinct phosphorylation pathways for activation of MAP kinase. 215 96

L-selectin is a leukocyte cell surface glycoprotein involved in carbohydrate-specific ligand binding which mediates rolling of leukocytes along endothelial surfaces. In addition to its role in adhesion, an intracellular signaling role for L-selectin has recently been recognized. In particular, cross-linking L-selectin leads to increased cytosolic Ca2+ levels and potentiation of the oxidative burst. As several cell surface glycoproteins have been shown to be linked to tyrosine kinases, we examined the hypothesis that L-selectin may be linked to pathways involving tyrosine phosphorylation in human neutrophils. Ligation of L-selectin by three different antibodies recognizing separate epitopes led to increased tyrosine phosphorylation of several cellular proteins as judged by anti-phosphotyrosine immunoblots of whole cell lysates with prominent bands at 40-42, 55-60, 70-72, and 105-120 kDa. The 42-kDa band comigrated with mitogen-activated protein (MAP) kinase as determined by immunoblotting with anti-MAP kinase antibody. This effect was specific for L-selectin, because antibodies against CD18, CD45, and CD10 did not increase tyrosine phosphorylation. Phosphorylation was not due to Fc binding, since F(ab')2 fragments of the anti-L-selectin antibodies were similarly effective, and the response was unaffected by Fc receptor blockade. Cross-linking of L-selectin was not required for enhanced tyrosine phosphorylation, because monovalent Fab fragments also increased tyrosine phosphorylation. The response to L-selectin antibodies was not inhibited by cytochalasin, suggesting that reorganization of the actin cytoskeleton was not required for this response. Sulfatides, sulfated glycolipids which may be natural ligands for L-selectin, also induced a rapid, dose-dependent increase in tyrosine phosphorylation. In addition, sulfatides, but not control glycolipids, resulted in enhanced tyrosine phosphorylation of MAP kinase. Both sulfatides and anti-L-selectin antibodies increased kinase activity of MAP kinase as determined by gel renaturation assay. The tyrosine kinase inhibitor, genistein, blocked the transient increase in intracellular Ca2+ and the oxidative burst induced by sulfatides, suggesting that this tyrosine phosphorylation is functionally important. We conclude that L-selectin is able to transmit intracellular signals, including increased tyrosine phosphorylation and activation of MAP kinase in neutrophils. We speculate that these events may contribute to the activation of neutrophils during adhesion.
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PMID:Signaling functions of L-selectin. Enhancement of tyrosine phosphorylation and activation of MAP kinase. 754 Oct 41

In addition to their role in bacterial killing, reactive oxygen intermediates (ROI) produced by the NADPH oxidase may participate in the regulation of intracellular pathways. We have recently demonstrated that ROI produced by the oxidase regulate tyrosine phosphorylation in neutrophils, possibly by alterations in the cellular redox state. The purpose of the present study was to characterize the identities of certain of the redox-sensitive tyrosine-phosphorylated substrates and the significance of the increased phosphorylation. As a prominent 42-44-kDa phosphorylated band was noted in oxidant-treated cells, we investigated the possible phosphorylation and activation of mitogen-activated protein (MAP) kinase under these conditions. Immunoprecipitation of MAP kinase followed by immunoblotting with anti-phosphotyrosine antibodies indicated that a 42-44-kDa polypeptide was tyrosine-phosphorylated in response to treatment of cells, either with the oxidizing agent diamide or with H2O2 in cells where catalase was inhibited. Using an in vitro renaturation assay with myelin basic protein as the substrate, oxidant-induced stimulation of kinase activity of a 42-44-kDa band was observed in both whole cell extracts and in MAP kinase immunoprecipitates. The mechanism of redox-sensitive activation of MAP kinase was examined. First, exposure of cells to oxidants caused a significant increase in the activity of MEK (the putative activator of MAP kinase), as determined by an in vitro kinase assay using recombinant catalytically inactive glutathione S-transferase-MAP kinase as the substrate. Additionally, oxidant treatment of cells resulted in inhibition of the activity of CD45, a protein tyrosine phosphatase known to dephosphorylate and inactivate MAP kinase. We conclude that oxidant treatment of neutrophils can activate MAP kinase by stimulating its tyrosine and (presumably) threonine phosphorylation via MEK activation, a response that may be potentiated by inhibition of MAP kinase dephosphorylation by phosphatases such as CD45.
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PMID:Activation of the mitogen-activated protein kinase signaling pathway in neutrophils. Role of oxidants. 798 67

Two myelin basic protein kinases designated MBPK-1 and MBPK-2 were purified to apparent homogeneity from extracts of bovine kidney cortex. The purified preparations exhibited an apparent M(r) approximately 40,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and approximately 42,000 (MBPK-1) and 45,000 (MBPK-2) by gel permeation chromatography. Up to 0.4 and 1.8 mol of phosphoryl groups were incorporated per mol of MBPK-1 and MBPK-2, respectively, on threonines following incubation with ATP. Autophosphorylation, incubation with protein phosphatase 2A2 (PP2A2), CD45, or T-cell protein tyrosine phosphatase did not affect MBPK-1 activity. Autophosphorylation increased by about 3-fold MBPK-2 activity. This autophosphorylation and activation was reversed by PP2A2 but not by CD45 or T-cell protein tyrosine phosphatase. MBPK-1 and MBPK-2 displayed a positive reaction with an antibody to mitogen-activated protein kinase. Purified preparations of protamine kinase were activated by about 1.5-6-fold and, after inactivation with PP2A2, were reactivated by about 30% by MBPK-1 and MBPK-2. Activation and reactivation correlated with the incorporation, respectively, of 0.1-0.5 and 0.5 mol of phosphoryl groups/mol of the protamine kinase on serines. The results show that MBPK-1 and MBPK-2 are protamine kinase-activating kinases and suggest that MBPK-1 and MBPK-2 may be related to mitogen-activated protein kinase.
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PMID:Phosphorylation and activation of protamine kinase by two forms of a myelin basic protein kinase from extracts of bovine kidney cortex. 839 73

T lymphocytes require two signals for activation. Recognition of antigen/MHC complexes by the T cell receptor delivers the first signal, while a second signal, delivered by the cell surface receptors CD80 and/or CD86 binding to the T cell surface molecule CD28, has been shown to be effective for the initiation of effective T cell responses. While some of the cytoplasmic effector molecules involved in T cell receptor signaling is known, little is known regarding those involved in the co-stimulation of T cells by CD28. Using the T cell leukemic cell line Jurkat as a model for T cell activation, we demonstrate that cross-linking CD28 using monoclonal antibodies causes tyrosine phosphorylation and activation of MAP kinase/ERK. This activation was rapid, peaking at approximately 5 minutes post CD28 cross-linking, and transient. Activation of MAP kinase/ERK occurred 3 fold less efficiently in a Jurkat line lacking functional p56lck (JCAM.1), and was almost undetectable in a line lacking CD45 (J45.01). These results suggest that CD28 cross-linking can activate intracellular signaling pathways via several different tyrosine kinases. Thus CD28 signaling can activate src family kinases lck and fyn, as well as the Tec family kinase emt/itk. Activation of any one or a combination of these tyrosine kinases may be sufficient for the activation of MAPK following CD28 cross-linking. Activation of MAPK has been shown to cause activation of AP-1 and other transcription factors via serine and/or threonine phosphorylation.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Activation of extracellular signal-regulated protein kinase (ERK/MAP kinase) following CD28 cross-linking: activation in cells lacking p56lck. 852 74

Overexpression of the transmembrane protein-tyrosine phosphatase (PTPase) CD45 in nonhematopoietic cells results in decreased signaling through growth factor receptor tyrosine kinases. Consistent with these data, insulin receptor signaling is increased when the CD45-related PTPase LAR is reduced by antisense suppression in a rat hepatoma cell line. To test whether the hematopoietic cell-specific PTPase CD45 functions in a manner similar to LAR by negatively modulating insulin receptor signaling in hematopoietic cells, the insulin-responsive human multiple myeloma cell line U266 was isolated into two subpopulations that differed in CD45 expression. In CD45 nonexpressing (CD45-) cells, insulin receptor autophosphorylation was increased by 3-fold after insulin treatment when compared to CD45 expressing (CD45+) cells. This increase in receptor autophosphorylation was associated with similar increases in insulin-dependent tyrosine kinase activation. These receptor level effects were paralleled by postreceptor responses. Insulin-dependent tyrosine phosphorylation of insulin receptor substrate 1 (IRS-1) and Shc was 3-fold greater in CD45- cells. In addition, insulin-dependent IRS-1/phosphatidylinositol 3-kinase association and MAP kinase activation in CD45- cells were also 3-fold larger. While expression of CD45 was associated with a decrease in the responsiveness of early insulin receptor signaling, interleukin 6-dependent activation of mitogen-activated protein kinase kinase and mitogen-activated protein kinase was equivalent between CD45- and CD45+ cells. These observations indicate that CD45 can function as a negative modulator of growth factor receptor tyrosine kinases in addition to its well-established role as an activator of src family tyrosine kinases.
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PMID:The transmembrane protein-tyrosine phosphatase CD45 is associated with decreased insulin receptor signaling. 855 83


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