Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.24 (mitogen-activated protein kinase)
 95,810 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The mitogenic neuropeptides bombesin and vasopressin markedly increased tyrosine and serine phosphorylation of multiple substrates in quiescent Swiss 3T3 fibroblasts, including two major bands of Mr 90,000 and 115,000. Tyrosine phosphorylation of these proteins was increased as judged by immunoprecipitation of 32Pi-labeled cells and immunoblotting of unlabeled cells with monoclonal antiphosphotyrosine antibodies, elution with phenyl phosphate, and phospho amino acid analysis. Phosphotyrosyl proteins generated by bombesin and vasopressin did not correspond either by apparent molecular weight or by immunological and biochemical criteria to several known tyrosine kinase substrates, including phospholipase C gamma, the microtubule-associated protein 2 kinase, GTPase-activating protein, or phosphatidylinositol kinase. The effect was rapid (within seconds), concentration dependent, and inhibited by specific receptor antagonists for both bombesin and vasopressin. The endothelin-related peptide, vasoactive intestinal contractor, also elicited a rapid and concentration-dependent tyrosine/serine phosphorylation of a similar set of substrates. These results demonstrate that neuropeptides, acting through receptors linked to GTP-binding proteins, stimulate tyrosine phosphorylation of a common set of substrates in quiescent Swiss 3T3 cells and suggest the existence of an additional signal transduction pathway in neuropeptide-induced mitogenesis.
 ...
PMID:Bombesin, vasopressin, and endothelin rapidly stimulate tyrosine phosphorylation in intact Swiss 3T3 cells. 164 10

In Swiss 3T3 fibroblasts a peptide mitogen bombesin, which acts through the phospholipase C-protein kinase C signaling pathway, stimulates DNA synthesis in a manner strictly dependent on the medium calcium concentration: [3H]thymidine incorporation into DNA in the presence of a saturating concentration of bombesin (10(-8) M) is 4-fold greater at 3.0 mM extracellular calcium as compared with a value obtained at 0.03 mM calcium. In the present study we attempted to identify the site and the mechanism of action of Ca2+ influx along the bombesin-induced mitogenic signaling pathway, by comparing bombesin effects at 0.03 and 3.0 mM of medium calcium. Bombesin induces the same extent of increases in [3H]inositol phosphates after 1 min, and comparable sustained increases in the cellular content of 1,2-diacylglycerol for up to 4 h, at either 0.03 or 3.0 mM calcium. Bombesin induces the same extent of phosphorylation of MARCKS protein, the major cellular substrate for protein kinase C, irrespective of the medium calcium concentration for at least 4 h. Moreover, diverse cellular responses elicited by bombesin, including c-fos expression, activation of microtubule-associated protein 2 kinase and S6 kinase, glucose uptake, and protein synthesis but not the release of arachidonic acid and its metabolites, are induced similarly at either 0.03 or 3.0 mM calcium. Down-regulation of cellular protein kinase C nearly completely abolishes bombesin effects on c-fos expression, S6 kinase activation, glucose uptake, and DNA synthesis. These results suggest that the target of Ca2+ influx in bombesin-induced mitogenic signaling pathway is not located along the phospholipase C-protein kinase C signal transduction system including cellular events in early G1 phase that exist downstream to protein kinase C action.
 ...
PMID:Role of Ca2+ influx in bombesin-induced mitogenesis in Swiss 3T3 fibroblasts. 184 53

We have investigated whether mitogen-activated protein (MAP) kinase cascade is essential for sustained contraction of smooth muscle cells of the rabbit rectosigmoid. We have identified MAP kinase as one of the enzymes activated by bombesin, performed immunologic studies blocking the activation of MAP kinase, and conducted confocal localization of MAP kinase in relation to heat-shock protein (HSP27), postulated to be involved in the sustained contraction of smooth muscle. Immunoblotting revealed two forms of MAP kinase (42 and 44 kDa). Activation of MAP kinase by bombesin was rapid, reaching a maximum in 30 s and subsequently declining. [D-Phe6,Leu13,psi(CH2NH),Phe14]BN-(6-14), a potent bombesin antagonist, and protein kinase C (PKC) inhibitors 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine, calphostin C, and chelerythrine inhibited the increase in MAP kinase induced by bombesin. Immunofluorescent dual labeling and confocal microscopy indicate that these two proteins are closely distributed in resting cells and that during bombesin-induced contraction MAP kinase translocates accompanied by HSP27. In conclusion, a series of events involving PKC activation, MAP kinase activation, and MAP kinase-HSP27 translocation could be the signaling pathway involved in bombesin-induced sustained contraction.
 ...
PMID:Activation of MAP kinase and translocation with HSP27 in bombesin-induced contraction of rectosigmoid smooth muscle. 749 59

The data presented in this paper demonstrate a new substance P (SP) binding site that is expressed on human monocytes. The apparent dissociation constant (Kd) for binding of 125I-labeled Bolton Hunter-SP (125I-BH-SP) to the receptor on monocyte membranes is 2.24 +/- 0.9 x 10(-7) M and the maximum binding capacity (Bmax) is 4.7 +/- 0.5 pmol/mg membrane protein. It could be excluded that this receptor is one of the known neurokinin (NK) type of receptors on the basis of binding characteristics for NK1, NK2, and NK3 agonists. Moreover, we demonstrate that the binding site is neither the bombesin receptor nor the serpin enzyme complex receptor nor the FMLP receptor. The order of potency for inhibition of 125I-BH-SP binding to the receptor on monocyte membranes is NK1 antagonist [D-Pro2,D-Trp7,9]SP > SP > NK3 agonist [MePhe7]SP > bombesin. Cross-linking studies with disuccinimidylsuberate, followed by SDS-PAGE analysis, revealed that 125I-BH-SP is specifically bound to a membrane protein with an apparent molecular mass of 47 kDa. At a functional level, SP induces the activation of MAP kinase in human monocytes. The ED50 for activation of MAP kinase positively correlated (r = 0.999, p < 0.0005) with the apparent affinity of the ligands applied in the 125I-BH-SP displacement studies. From these results, we conclude that this SP binding site on monocytes is a non-NK receptor protein that is functionally linked to the activation of MAP kinase.
 ...
PMID:Monocytes express a non-neurokinin substance P receptor that is functionally coupled to MAP kinase. 751 35

Mitogenic stimulation of Swiss 3T3 fibroblasts with bombesin results in receptor-mediated activation of a complex array of effectors, including phospholipase C beta and mitogen-activated protein (MAP) kinase. Incubation of Swiss 3T3 fibroblasts with the 11-amino acid [D-Arg1,D-Phe5,D-Trp7,9,Leu11]substance P peptide inhibited bombesin-stimulated cell proliferation and phospholipase C beta activation even at high bombesin concentrations. The peptide did not inhibit the activation of phospholipase C beta by a GTPase-deficient form of the Gq-like protein, G16, indicating that the peptide does not inhibit phospholipase C beta and is acting at a point upstream of the activated form of the G protein alpha subunit. The peptide inhibited MAP kinase activation at low bombesin concentrations, but unlike phospholipase C beta, this inhibition could be overcome with 30 nM bombesin. In control Swiss 3T3 cells, bombesin did not measurably activate Ras or Raf-1 above basal levels. Following incubation of the cells with the [D-Arg1,D-Phe5,D-Trp7,9,Leu11]substance P peptide, 50 nM bombesin activated Raf-1 4-6-fold over basal levels. Platelet-derived growth factor-stimulated activities of PLC, Ras, Raf-1, and MAP kinase were unaltered after incubation of Swiss 3T3 cells with the [D-Arg1,D-Phe5,D-Trp7,9,Leu11]substance P peptide, as was platelet-derived growth factor-stimulated growth of the Swiss 3T3 cells. Thus, the peptide behaves as an antagonist that differentially inhibited phospholipase C beta and MAP kinase signal transduction pathways. The growth arrest observed with the peptide indicates that the bombesin-stimulated activation of MAP kinase is not sufficient to support mitogenesis in Swiss 3T3 cells.
 ...
PMID:Differential modulation of bombesin-stimulated phospholipase C beta and mitogen-activated protein kinase activity by [D-Arg1,D-Phe5,D-Trp7,9,Leu11]substance P. 753 38

Sphingosylphosphorylcholine (SPC) is a potent mitogen for Swiss 3T3 cells, but the signaling mechanisms involved are poorly characterized. Here, we report that addition of SPC induces a rapid and transient activation of p42 mitogen-activated protein kinase (p42MAPK) in these cells. SPC-induced p42MAPK activation peaked at 5 min and was undetectable after 30 min of incubation with SPC. The effect of SPC on p42MAPK activation was comparable to that induced by bombesin and platelet-derived growth factor. As SPC strongly induced phosphorylation of the major protein kinase C (PKC) substrate 80K/MARCKS in either intact or permeabilized cells, we examined whether PKC could be involved in SPC-induced p42MAPK activation. Here, we demonstrate that p42MAPK activation by SPC was dependent on PKC activity as shown by inhibition of PKC with the bisindolymaleimide GF 109203X or down-regulation of PKC by prolonged treatment of Swiss 3T3 cells with phorbol esters. Activation of both PKC and p42MAPK by SPC was markedly inhibited by treatment with pertussis toxin, implicating a G proteins(s) of the Gi/G(o) subfamily in the action of SPC. SPC-induced rapid activation of a downstream target of p42MAPK, p90 ribosomal S6 kinase (p90rsk), also required PKC and a pertussis toxin-sensitive G protein. In addition, SPC-induced mitogenesis was dependent on a Gi protein in Swiss 3T3 cells. SPC also induced p42MAPK activation and DNA synthesis in secondary cultures of mouse embryo fibroblasts through a pertussis toxin-sensitive pathway. As G proteins link many cell surface receptors to effector proteins, we hypothesize, therefore, that SPC could bind to a receptor that mediates at least some of its biological effects in Swiss 3T3 cells and mouse embryo fibroblasts.
 ...
PMID:Sphingosylphosphorylcholine activation of mitogen-activated protein kinase in Swiss 3T3 cells requires protein kinase C and a pertussis toxin-sensitive G protein. 759 45

Cholecystokinin (CCK) has recently been shown to activate mitogen-activated protein (MAP) kinase in rat pancreatic acini [Duan and Williams, Am. J. Physiol. 267 (Gastrointest. Liver Physiol. 30): G401-G408, 1994]. To evaluate the mechanism of MAP kinase activation, we studied the effects of CCK on MAP kinase kinase (MEK) in rat pancreatic acini. Two forms of MEK were identified by immunoblotting, using antibodies specific to MEK1 and MEK2. MEK activity in acinar extracts and after immunoprecipitation with anti-MEK was detected using a recombinant fusion protein, glutathione S-transferase-MAP kinase, as a substrate. MEK activity rapidly increased after stimulation of acini by CCK, with significant stimulation at 1 min and a maximal effect at 5 min, followed by a slow decline to slightly above control levels after 30 min. The threshold concentration of CCK was approximately 10 pM, and the maximal effect was induced by 1 nM CCK, which increased MEK activity by 120%. In addition to CCK, bombesin and carbachol, but not secretin or vasoactive intestinal peptide, enhanced MEK activity. Phorbol ester mimicked the effect of CCK, whereas ionomycin and thapsigargin failed to activate MEK. We further studied the activation of Ras, an important component leading to activation of MEK by growth factors. Ras in acini was immunoprecipitated and identified by Western blotting. CCK and 12-O-tetradecanoylphorbol-13-acetate stimulated the incorporation of GTP into Ras, a requirement for its activation, reaching a maximum at 10 min of approximately 120% over control. In conclusion, the activation of MAP kinase by CCK can be explained by activation of MEK and may involve the activation of Ras by a protein kinase C-dependent mechanism.
 ...
PMID:Activation of MAP kinase kinase (MEK) and Ras by cholecystokinin in rat pancreatic acini. 761 6

Elevation of intracellular cAMP by forskolin, 8-bromoadenosine 3':5'-cyclic monophosphate, and prostaglandin E1, in synergy with insulin, stimulated DNA synthesis in quiescent Swiss 3T3 cells to the same level achieved by platelet-derived growth factor (PDGF) or bombesin. Both forskolin and 8-bromoadenosine 3':5'-cyclic monophosphate stimulated a significant increase in cell number which, in the presence of insulin, reached the same levels achieved with PDGF. Treatment with either PDGF or bombesin caused a marked and persistent stimulation of p42MAPK and p44MAPK. In striking contrast, no activation was seen with mitogenic combinations of cAMP as shown by three different assays. Swiss 3T3 cells stably transfected with a constitutively activated Gs alpha subunit were 100-fold more sensitive to the mitogenic effects of forskolin but in this distinct cellular model forskolin did not activate p42MAPK. Swiss 3T3 cells stably transfected with interfering mutants of MEK-1 showed a 60% decrease in PDGF-stimulated p42 MAPK activation, but there was no inhibition of the mitogenic effect of forskolin in these cells. Furthermore, the upstream kinases MEK-1/MEK-2 and p74raf-1 were not activated by mitogenic combinations of cAMP while PDGF caused marked stimulation of their activity. Treatment of 3T3 cells with forskolin attenuated PDGF-stimulated p74raf-1 and p42MAPK activation but enhanced the mitogenic effects of this agent. Mitogenic combinations of cAMP strongly stimulated the phosphorylation and activation of p70s6k an effect that was inhibited by rapamycin. This agent markedly inhibited cAMP-stimulated DNA synthesis suggesting a critical role for p70s6k in cAMP mitogenic signaling. These results demonstrate that cAMP-induced mitogenesis can be dissociated from activation of the mitogen-activated protein kinase cascade and that this is not an obligatory point of convergence in mitogenic signaling in Swiss 3T3 cells.
 ...
PMID:Dissociation of cAMP-stimulated mitogenesis from activation of the mitogen-activated protein kinase cascade in Swiss 3T3 cells. 767 77

The specific inhibitors of the endoplasmic reticulum Ca2+ pump, thapsigargin and 2,5-di-tert-butylhydroquinone (DBHQ), stimulated reinitiation of DNA synthesis in synergy with either phorbol 12,13-dibutyrate or bombesin in Swiss 3T3 cells. Maximum stimulation was achieved at 0.5 nM thapsigargin and 7.5 microM DBHQ. Kinetics of [3H]thymidine incorporation were consistent with exit from G0 and entry into S phase. Autoradiography of labeled nuclei showed that the increase in [3H]thymidine incorporation was due to an increase in the proportion of cells entering into DNA synthesis. Down-regulation or selective inhibition of protein kinase C abolished this synergistic stimulation of DNA synthesis. Thapsigargin and DBHQ did not potentiate protein kinase C-mediated signals such as direct phosphorylation of myristoylated alanine-rich C-kinase substrate, activation of mitogen-activated protein kinase, and tyrosine phosphorylation of bands 110,000-130,000 and 70,000-80,000. Thapsigargin and DBHQ caused a marked reduction in the ability of bombesin to induce a rapid and transient increase in intracellular Ca2+ via depletion of total cellular Ca2+, measured by 45Ca2+ content. The synergistic stimulation of DNA synthesis by DBHQ and phorbol 12,13-dibutyrate was dependent on a high concentration of extracellular Ca2+ (ED50 = 410 microM) and was preferentially inhibited by the inhibitor of Ca2+ influx econozole. This suggests a role for Ca2+ entry in growth control. This is the first time that either thapsigargin or DBHQ has been shown to stimulate the reinitiation of DNA synthesis in any target cell.
 ...
PMID:Thapsigargin and di-tert-butylhydroquinone induce synergistic stimulation of DNA synthesis with phorbol ester and bombesin in Swiss 3T3 cells. 779 54

Both bombesin and epidermal growth factor (EGF) are potent mitogens in Swiss 3T3 cells that nonetheless have dissimilar receptor structures. To explore possible common intracellular events involved in the stimulation of cellular growth by these two peptides, we have evaluated the regulation of the mitogen-activated protein (MAP) kinase. Exposure of Swiss 3T3 cells to bombesin, EGF or the protein kinase C activator phorbol 12-myristate 13-acetate (PMA) causes the rapid and transient stimulation of the enzyme activity. Pretreatment of cells with the protein kinase inhibitor H-7, or down-regulation of cellular protein kinase C by prolonged exposure to PMA, causes a decrease of over 90% in the activation of MAP kinase by bombesin. In contrast, these treatments have no effect on the stimulation of MAP kinase by EGF. The stimulation of MAP kinase activity by bombesin is dose-dependent, occurring over a narrow concentration range of the peptide. Both EGF and bombesin stimulate the phosphorylation of an immunoprecipitable MAP kinase protein migrating at 42 kDa on SDS/PAGE. Phosphoamino acid analysis of this phosphorylated protein reveals that EGF and bombesin stimulate phosphorylation on tyrosine, threonine and serine residues. Tyrosine phosphorylation of the enzyme, as evaluated by antiphosphotyrosine blotting of the immunoprecipitated protein, reveals that the time course of phosphorylation by both mitogens correlates with stimulation of enzyme activity. These results provide further evidence for the convergence of discrete pathways emanating from tyrosine kinase and G-protein-linked receptors in the regulation of MAP kinase.
 ...
PMID:Bombesin and epidermal growth factor stimulate the mitogen-activated protein kinase through different pathways in Swiss 3T3 cells. 838 Sep 87


1 2 3 4 5 6 7 8 Next >>