UNIPROT:P51812 - FACTA Search

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Query: UNIPROT:P51812 (mitogen-activated protein)
10,636 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Son of sevenless-1 and -2 (Sos-1 and -2) are guanosine nucleotide exchange factors implicated in the activation of Ras by both the insulin and epidermal growth factor signal transduction pathways. Ras appears to function by initiating the activation of cellular protein kinases including mitogen-activated protein (MAP) kinases. Sos proteins contain numerous sequences in their carboxyl-terminal regions which correspond to consensus sites for MAP kinase phosphorylation. To examine whether these sites are substrates for MAP kinases, the cDNA encoding Drosophila Sos (dSos) was tagged with sequences encoding the major antigenic epitope of the influenza virus hemagglutinin (HA) to create a dSosHA fusion construct. dSosHA was transiently expressed in COS-1 cells and immunoprecipitated with anti-HA antibodies. When immune complexes were incubated with purified MAP kinase and [gamma-32P]ATP, a phosphorylated band of 180 kDa was observed when analyzed by SDS-polyacrylamide gel electrophoresis. This band was not present in immunoprecipitations from cells transfected with vector alone. No phosphorylation of the 180 kDa band was seen when immunoprecipitates were incubated with [gamma-32P]ATP in the absence of MAP kinase. Two dimensional analysis of tryptic peptides from dSosHA phosphorylated by MAP kinase in vitro revealed two major phosphorylated species that were also found in dSosHA isolated from COS-1 cells labeled with 32Pi. These results are consistent with the hypothesis that a feedback loop exists wherein growth factor-activated MAP kinases phosphorylate and regulate Sos proteins.
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PMID:Phosphorylation of the Ras nucleotide exchange factor son of sevenless by mitogen-activated protein kinase. 810 39

Treatment of Chinese hamster ovary (CHO) cells over-expressing the human insulin receptor (CHO-HIRc) with the insulin mimetic agent, vanadate, resulted in a dose- and time-dependent tyrosine phosphorylation of two proteins with apparent molecular sizes of 42 kDa (p42) and 44 kDa (p44). However, vanadate was unable to stimulate the tyrosyl phosphorylation of the beta-subunit of the insulin receptor. By using myelin basic protein (MBP) as the substrate to measure mitogen-activated protein (MAP) kinase activity in whole cell lysates, vanadate-stimulated tyrosyl phosphorylation of p42 and p44 was associated with a dose- and time-dependent activation of MAP kinase activity. Furthermore, affinity purification of cell lysates on anti-phosphotyrosine agarose column followed by immunoblotting with a specific antibody to MAP kinases demonstrated that vanadate treatment increased the tyrosyl phosphorylation of both p44mapk and p42mapk by several folds, as compared to controls, in concert with MAP kinase activation. In addition, retardation in gel mobility further confirmed that vanadate treatment increased the phosphorylation of p44mapk and p42mapk in CHO-HIRc. A similar effect of vanadate on MAP kinase tyrosyl phosphorylation and activation was also observed in CHO cells over-expressing a protein tyrosine kinase-deficient insulin receptor (CHO-1018). These results demonstrate that the protein tyrosine kinase activity of the insulin receptor may not be required in the signaling pathways leading to the vanadate-mediated tyrosyl phosphorylation and activation of MAP kinases.
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PMID:Activation of mitogen activated protein (MAP) kinases by vanadate is independent of insulin receptor autophosphorylation. 813 57

Directed migration or chemotaxis of arterial smooth muscle cells (SMC) contributes to intimal SMC accumulation, a key event in the development of atherosclerotic lesions and in restenosis after angioplasty. The present study compares and contrasts insulin-like growth factor I (IGF-I) and platelet-derived growth factor (PDGF-BB) as chemoattractants and mitogens for human arterial SMC. Compared with PDGF-BB, IGF-I is a weaker SMC mitogen. Thus, PDGF-BB, but not IGF-I, evokes a strong and rapid activation of mitogen-activated protein (MAP) kinase kinase and MAP kinase. However, IGF-I is a potent stimulator of directed migration of human arterial SMC, as measured in a Boyden chamber assay. The half-maximal concentration for migration is similar to the Kd for IGF-I receptor interaction. An IGF-I receptor-blocking antibody blocks the effects of IGF-I, IGF-II, and insulin, indicating that the effects are indeed mediated through the IGF-I receptor. The maximal effect of IGF-I on directed migration ranges between 50% and 100% of the effect of PDGF-BB, the strongest known chemoattractant for SMC. The ability of IGF-I and PDGF-BB to induce chemotaxis coincides with their ability to stimulate phosphatidylinositol turnover, diacylglycerol formation, and intracellular Ca2+ flux and suggests that these signaling pathways, but not activation of the MAP kinase cascade, are required for chemotaxis of human arterial SMC.
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PMID:Insulin-like growth factor-I and platelet-derived growth factor-BB induce directed migration of human arterial smooth muscle cells via signaling pathways that are distinct from those of proliferation. 813 65

Interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) have previously been reported to induce rapid phosphorylation of the mitogen-activated protein (MAP) kinase. However, little is known about signaling events initiated by both hematopoietins that occur downstream of the MAP kinase. MAP kinase has been shown to phosphorylate the AP-1 transcription factor and also to activate two kinases designated insulin-stimulated protein kinase-1 and MAP kinase-activated protein (MAP-KAP) kinase 2. We show here that IL-3 and GM-CSF induce MAPKAP kinase 2 activity in the human megakaryoblastic leukemia cell line MO7 and phosphorylate the human small heat shock protein Hsp 27 on serine residues in vitro. GM-CSF also induced Hsp 27 phosphorylation in neutrophils in a range similar to that observed in MO7 cells, suggesting that MAPKAP kinase 2-mediated Hsp 27 activation occurs independently of proliferation. Hsp 27 phosphorylation was dose-dependent, occurred as early as 5 minutes after factor exposure, and was inhibited by the tyrosine kinase inhibitors genistein and herbimycin A. Furthermore, the protein phosphatase A2 abolished IL-3- and GM-CSF-induced serine phosphorylation of Hsp 27. Taken together, our findings indicate that tyrosine phosphorylation of MAP kinase is a prerequisite for serine phosphorylation of Hsp 27, which is mediated by MAPKAP kinase 2. Hsp 27 has shown activation-dependent translocation from the cytosolic to the nuclear region and has been linked to the cellular stress response. However, its precise function is largely unknown. Our data identify Hsp 27 as a target of the IL-3/GM-CSF stimulation pathway that involves MAP kinase and MAPKAP kinase 2. In addition, our results indicate that Hsp 27 may be target of phosphorylation events not only in the stress response but also in unstressed cells responding to cytokine stimulation.
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PMID:Interleukin-3 and granulocyte-macrophage colony-stimulating factor induce activation of the MAPKAP kinase 2 resulting in in vitro serine phosphorylation of the small heat shock protein (Hsp 27). 1101 49

Activation of mitogen-activated protein (MAP) kinase represents an important mechanism in hormonal regulation. To clarify the role of MAP kinase activation in insulin action, we compared the activation of the enzyme in Rat-1 cells transfected with wild-type (Hirc) and mutant insulin receptors in which the 2 carboxyl-terminal tyrosines were substituted with phenylalanine (Y/F2). Expression of the Y/F2 mutant receptor enhanced the responsiveness of MAP kinase to insulin. Moreover, the insulin responsiveness of the activator of this enzyme, MAP kinase kinase, was also increased in these cells. To explore the early signaling events that might account for this increase in responsiveness, we evaluated the tyrosine phosphorylation of the insulin receptor substrate, IRS-1, and its subsequent association with phosphatidylinositol (PI)-3 kinase. In both cell types, insulin led to a dose-dependent increase in the association of tyrosine phosphorylated IRS-1 with the SH2 domain of the p85 regulatory subunit of PI-3 kinase, and also increased the amount of PI kinase activity detected in anti-IRS-1 immunoprecipitates. The effect of insulin was significantly greater in Y/F2 cells, as determined in both assays. In previous studies, cells bearing this receptor mutant exhibited an identical metabolic response but enhanced mitogenic response to insulin when compared with wild-type receptor. These data provide further evidence for divergence of the mitogenic and metabolic signaling pathways at or near the insulin receptor.
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PMID:Mutation of the two carboxyl-terminal tyrosines in the insulin receptor results in enhanced activation of mitogen-activated protein kinase. 814 49

GRB-2 is a small SH2- and SH3 domain-containing adapter protein that associates with the mammalian SOS homolog to regulate p21ras during growth factor signaling. During insulin stimulation, GRB-2 binds to the phosphorylated Y895VNI motif of IRS-1. Substitution of Tyr-895 with phenylalanine (IRS-1F-895) prevented the IRS-1-GRB-2 association in vivo and in vitro. The myeloid progenitor cell line, 32-D, is insensitive to insulin because it contains few insulin receptors and no IRS-1. Coexpression of IRS-1 or IRS-1F-895 with the insulin receptor was required for insulin-stimulated mitogenesis in 32-D cells, while expression of the insulin receptor alone was sufficient to mediate insulin-stimulated tyrosine phosphorylation of Shc and activation of p21ras and mitogen-activated protein (MAP) kinase. The Shc-GRB-2 complex formed during insulin stimulation is a possible mediator of p21ras and MAP kinase activation in IRS-1-deficient 32-D cells. Interestingly, IRS-1, but not IRS-1F-895, enhanced the stimulation of MAP kinase by insulin in 32-D cells expressing insulin receptors. Thus, IRS-1 contributes to the stimulation of MAP kinase by insulin, probably through formation of the IRS-1-GRB-2 complex at Tyr-895. Our results suggest that the Shc-GRB-2 complex and the activation of p21ras-dependent signaling pathways, including MAP kinase, are insufficient for insulin-stimulated mitogenesis and that the essential function(s) of IRS-1 in proliferative signaling is largely unrelated to IRS-1-GRB-2 complex formation.
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PMID:Role of IRS-1-GRB-2 complexes in insulin signaling. 819 3

The role of mitogen-activated protein (MAP) kinase in the regulation of glucose metabolism has been investigated by comparing the effects of insulin and epidermal growth factor (EGF) on MAP kinase activation, glucose transport, and glycogen synthase in 3T3-L1 adipocytes. Insulin or EGF treatment for 5 min increased p42mapk and p44mapk activity to the same extent as determined by myelin basic protein kinase activity measurements and phosphotyrosine immunoblotting. The profiles of myelin basic protein kinase activity following MonoQ chromatography of extracts obtained from cells incubated with insulin or EGF were almost identical. Insulin increased glucose transport and GLUT4 translocation to the cell surface by 15- and 7-fold, respectively. EGF had no significant effect on these processes. Insulin increased the glycogen synthase ratio (-Glc-6-P/+Glc-6-P) by 7.5- and 3.5-fold in the presence and absence of glucose, respectively. EGF increased the ratios by only 2- and 1.3-fold, respectively. EGF did not appear to inhibit downstream of MAP kinase, because when adipocytes were incubated with insulin plus EGF, the stimulation of glucose transport and glycogen synthase was similar to that observed with insulin alone. These findings indicate that activation of the MAP kinase isoforms p42mapk and p44mapk is not sufficient for the activation of glucose transport and glycogen synthase in 3T3-L1 adipocytes.
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PMID:Mitogen-activated protein kinase activation is not sufficient for stimulation of glucose transport or glycogen synthase in 3T3-L1 adipocytes. 825 68

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

Perturbations of fetal growth produce parallel but disproportionate changes in fetal liver growth that correlate with circulating fetal insulin concentration. We have studied the effects of insulin and two hepatotrophic factors, transforming growth factor-alpha (TGF alpha) and hepatocyte growth factor (HGF), on DNA synthesis by fetal and adult rat hepatocytes in primary culture. Using serum-free Minimum Essential Medium, fetal hepatocytes synthesized DNA without growth factors, unlike adult hepatocytes. Insulin augmented fetal hepatocyte DNA synthesis after 16-24 h in culture. In contrast, TGF alpha or HGF maximally stimulated fetal hepatocyte DNA synthesis after 40 h in culture. Insulin and TGF alpha were not synergistic in stimulating fetal hepatocyte DNA synthesis, but were synergistic in their action on adult hepatocytes. Brief (10-min) exposure of fetal hepatocytes to TGF alpha or HGF, but not insulin, activated mitogen-activated protein kinases 4-fold. Prolonged (24-h) exposure to TGF alpha or HGF abolished the ability of either to activate mitogen-activated protein kinases, whereas insulin had no effect. Maternal fasting for 48 h before isolation and culturing of fetal hepatocytes abolished the in vitro stimulation of DNA synthesis by insulin without affecting TGF alpha action. We conclude that insulin has growth-promoting actions on fetal hepatocytes that are distinct and independent from those of TGF alpha of HGF.
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PMID:Evidence for a direct hepatotrophic role for insulin in the fetal rat: implications for the impaired hepatic growth seen in fetal growth retardation. 829 72

Insulin-induced activation of extracellular signal-regulated kinases [ERKs, also known as mitogen-activated protein (MAP) kinases] is mediated by Ras. Insulin activates Ras primarily by increasing the rate of guanine nucleotide-releasing activity. Here, we show that insulin-induced activation of ERKs was enhanced by stable overexpression of growth factor receptor-bound protein 2 (GRB2) but not by overexpression of GRB2 proteins with point mutations in the Src homology 2 and 3 domains. Moreover, a dominant negative form of Ras (with Ser17 substituted with Asn) blocked insulin-induced activation of ERKs in cells that overexpressed GRB2. GRB2 overexpression led to increased formation of a complex between the guanine nucleotide-releasing factor Sos (the product of the mammalian homolog of son of sevenless gene) and GRB2. In response to insulin stimulation, this complex bound to tyrosine-phosphorylated IRS-1 (insulin receptor substrate-1) and Shc. In contrast to the activated epidermal growth factor receptor that binds the GRB2-Sos complex directly, activation of the insulin receptor results in the interaction of GRB2-Sos with IRS-1 and Shc, thus linking the insulin receptor to Ras signaling pathways.
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PMID:The function of GRB2 in linking the insulin receptor to Ras signaling pathways. 831 35

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