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

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.
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PMID:Activation of MAP kinase and translocation with HSP27 in bombesin-induced contraction of rectosigmoid smooth muscle. 749 59

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.
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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

Treatment of fibroblasts with wortmannin or demethoxyviridin, two potent inhibitors of phosphatidylinositol 3-kinase, prevents the activation of ribosomal protein S6 kinase, which is induced by a variety of external stimuli. Concentrations giving 50% inhibition of 45 nM (wortmannin) and 400 nM (demethoxyviridin) were obtained when epidermal growth factor was used as an S6 kinase activator; with platelet-derived growth factor, the concentrations giving 50% inhibition were about three-times higher. Western-blot analysis and immunocomplex kinase assays showed that wortmannin and demethoxyviridin specifically block the phosphorylation and activation of p70 S6 kinase without affecting the M(r) 90,000 ribosomal S6 kinase (p90rsk) or mitogen-activated protein kinases. Consistent with the irreversible nature of the inhibition of phosphatidylinositol 3-kinase by these compounds, treatment of cells with wortmannin, followed by washing out of the inhibitor, still led to inhibition of p70 S6 kinase activation. Several S6 kinase agonists not previously known to activate phosphatidylinositol 3-kinase (A23187, bombesin and phorbol 12-myristate 13-acetate) were found to increase the production of phosphatidylinositol 3,4,5-trisphosphate in a wortmannin-sensitive manner. These results support a model in which phosphatidylinositol 3-kinase acts upstream of p70 S6 kinase in a mitogenic signalling pathway; the existence of a phosphatidylinositol 3-kinase-independent pathway is also evident.
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PMID:Selective inhibition of p70 S6 kinase activation by phosphatidylinositol 3-kinase inhibitors. 760 12

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.
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PMID:Activation of MAP kinase kinase (MEK) and Ras by cholecystokinin in rat pancreatic acini. 761 6

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.
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PMID:Bombesin and epidermal growth factor stimulate the mitogen-activated protein kinase through different pathways in Swiss 3T3 cells. 838 Sep 87

Stimulation of pancreatic acini from male Sprague-Dawley rats by both cholecystokinin (CCK)-8 and anisomycin caused an increase in p46jnk and p55jnk activities. Both forms of c-Jun amino-terminal kinase (JNK) were slightly activated at 5 min, reached a maximum at 30 min, and remained significantly increased at 60 min of CCK stimulation. By contrast, p42mapkwas activated fully by 5 min. In pancreatic acini stimulated with different concentrations of CCK for 30 min, the minimal and maximal JNK responses were observed at 30 pm and 100 nM CCK, respectively; p42mapk activation was, as previously reported, much more sensitive, with maximal activation by 1 nm CCK. Carbachol and bombesin also stimulated JNK activity, while vasoactive intestinal peptide did not. Neither activating protein kinase C nor increasing intracellular Ca2+ significantly activated JNK. In in vivo experiments, rats were infused intravenously for 5 and 15 min with a secretory (0.1 microg/kg/h) or supramaximal (10 microg/kg/h) dose of the CCK analog caerulein (CER). Secretory doses of CER induced a 4-fold increase of both forms of JNK in pancreatic tissue at 5 and 15 min, while at the same time points, supramaximal stimulation with CER caused 4- and 27-fold increases, respectively, of these kinase activities. The secretory dose of CER slightly increased the activities of both forms of mitogen-activated protein kinase, while the supramaximal dose induced a 10-fold increase of p42mapk at 5 min. In conclusion, JNKs and mitogen-activated protein kinases are rapidly activated in rat pancreatic acini stimulated with CCK as well as in pancreatic tissue during in vivo stimulation with CER. The large response to supramaximal CER stimulation may be of importance in the early pathogenesis of acute pancreatitis.
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PMID:Jun kinases are rapidly activated by cholecystokinin in rat pancreas both in vitro and in vivo. 862 33

The novel substance P (SP) analogue, [D-Arg1,D-Trp5,7,9,Leu11]SP like [D-Arg1,D-Phe5,D-Trp7,9,Leu11]SP inhibited DNA synthesis induced by bombesin, vasopressin, and bradykinin, but did not interfere with the mitogenic response induced by other growth factors or pharmacological agents in Swiss 3T3 cells. [D-Arg1,D-Trp5, 7,9,Leu11]SP reversibly inhibited bombesin-induced DNA synthesis, causing a 6-fold greater rightward shift in the bombesin dose response than [D-Arg1,D-Phe5,D-Trp7,9,Leu11]SP at identical concentrations (10 microM). We found that the new, more potent, SP analogue coordinately and reversibly inhibited bombesin-induced Ca2+ mobilization and protein kinase C (PKC) and mitogen-activated protein (MAP) kinase activation. The dose-response curves for bombesin-induced Ca2+ mobilization and MAP kinase activation were similarly displaced (51- and 40-fold, respectively) by [D-Arg1, D-Trp5,7,9,Leu11]SP. In addition, [D-Arg1,D-Trp5,7,9,Leu11]SP reversibly inhibited bombesin-induced tyrosine phosphorylation of Mr 110,000-130,000 and 70,000-80,000 bands as well as p125 focal adhesion kinase. [D-Arg1,D-Trp5,7,9,Leu11]SP also reversibly and coordinately inhibited vasopressin-induced Ca2+ mobilization, PKC stimulation, MAP kinase activation, tyrosine phosphorylation, and DNA synthesis in Swiss 3T3 cells. Surprisingly, deletion of the terminal Leu of [D-Arg1,D-Phe5,D-Trp7,9,Leu11]SP to yield [D-Arg1, D-Phe5,D-Trp7,9]SP1-10 resulted in a selective loss of inhibitory activity of this analogue against bombesin- but not vasopressin-stimulated DNA synthesis, Ca2+ mobilization, and MAP kinase activation. Collectively, these results suggest that SP analogues act at the receptor level to coordinately and reversibly antagonize bombesin- or vasopressin-induced signal transduction in Swiss 3T3 cells.
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PMID:[D-Arg1,D-Trp5,7,9,Leu11]Substance P coordinately and reversibly inhibits bombesin- and vasopressin-induced signal transduction pathways in Swiss 3T3 cells. 891 Jun 12

Functional bombesin receptors were identified in most human glioblastoma cell lines examined (approximately 85% of lines). Bombesin stimulated the release of intracellular Ca2+ in human adult (U-373MG, D-247MG, U-118MG, U-251MG, D-245MG, U-105MG, D-54MG, A-172MG, and D-270MG lines) and pediatric (SJ-S6 and SJ-G2 lines) glioblastoma cell lines. Stimulation of the glioblastoma cell line U-373MG with bombesin or gastrin-releasing peptide (GRP) induced mitogenesis, measured by [3H]thymidine incorporation into DNA, and stimulated the tyrosine phosphorylation of the mitogen-activated protein (MAP) kinases (Erk1 and Erk2). The stimulation of the MAP kinase phosphorylation in U-373MG cells was time- and peptide concentration-dependent. Both bombesin and GRP showed similar potencies in stimulation of intracellular Ca2+ release and activation of the MAP kinase pathway in U-373MG cells, whereas neuromedin B (NMB) peptide was less potent. Bombesin and GRP induced the release of cytosolic Ca2+ in a concentration-dependent manner. Because bombesin and GRP were more potent than NMB peptide in increasing the cytosolic Ca2+ levels in U-373MG cells, we concluded that the BB2 subtype (also known as GRP-preferring receptor subtype) of the bombesin receptor is expressed in this cell line. The bombesin receptor antagonist ([Leu13-psi(CH2NH)Leu14]bombesin) blocked bombesin induced Ca2+ release and attenuated MAP kinase activation in U-373MG cells demonstrating that bombesin is acting through a receptor-dependent mechanism. This study indicates that functional bombesin receptors are widely expressed in human glioblastoma cell lines.
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PMID:Functional expression of bombesin receptor in most adult and pediatric human glioblastoma cell lines; role in mitogenesis and in stimulating the mitogen-activated protein kinase pathway. 922 28

We have investigated the hypotheses that 1) bombesin activation of protein kinase C (PKC) results in the hydrolysis of sphingolipids and the production of ceramide and that 2) ceramide produced on activation by bombesin mediates sustained contraction of smooth muscle cells by activation of PKC and mitogen-activated protein (MAP) kinase. Ceramide production was assessed using a technique that involved benzoylation of purified ceramide extracts, followed by reverse-phase high-performance liquid chromatography. Contraction of smooth muscle cells isolated from the rabbit rectosigmoid and stimulated with bombesin gave a significant increase in newly formed ceramide (38 +/- 3.5%). 12-O-tetradecanoylphorbol-13-acetate also induced production of ceramide, which was blocked by calphostin C. The short-chain permeable C2 ceramide induced a sustained contraction and activation of MAP kinase, which was blocked by calphostin C. The increase in MAP kinase activity was maximal at 30 s and declined at 2 min. The data suggest that stimulation of smooth muscle cells by bombesin results in a functional coupling between sn-1,2-diacylglycerol (DAG)/ PKC and a sphingomyelinase, whereby DAG activates the hydrolysis of sphingomyelin to produce ceramide. Ceramide in turn activates PKC, which then activates MAP kinase. This could be the basis for the sustained contraction observed with bombesin.
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PMID:Bombesin-stimulated ceramide production and MAP kinase activation in rabbit rectosigmoid smooth muscle cells. 922

Bombesin has been reported to stimulate cholecystokinin (CCK) secretion from rat duodeno-jejunal I-cells. Bombesin was shown to activate mitogen-activated protein kinases (MAPKs) in cell types such as Swiss 3T3 fibroblasts and rat pancreatic acinar cells. No information is available on whether MAPK is activated in intestinal endocrine cells upon bombesin stimulation. This was studied by using the CCK-producing enteroendocrine cell line STC-1. Bombesin stimulated markedly and transiently both p42(MAPK) and p44(MAPK), with a maximum at 2 min, and a decrease to basal levels within 10 min. As expected, bombesin stimulated MAPK kinase 1 (MEK-1) activity. Activation of protein kinase C (PKC) with PMA also stimulated p42(MAPK), p44(MAPK) and MEK-1. Treatment of cells with PD 098059 (at 10 microM or 30 microM), which selectively inhibits MEK phosphorylation, blocked bombesin-induced p42(MAPK) and p44(MAPK) activation for at least 90 min. However, PD 098059 inhibited bombesin- and PMA-stimulated CCK secretion during the first 15 min, but failed to significantly reduce CCK release at later times. Inhibition of PKC with staurosporine, or PKC down-regulation by prolonged treatment with PMA, both drastically decreased MEK-1, p42(MAPK) and p44(MAPK) activation upon bombesin stimulation. Additionally, PKC activation appeared to be required for both MAPK-dependent (early) and -independent (late) CCK responses to bombesin. It is concluded that the early CCK secretory response of STC-1 cells to bombesin involves MAPK pathway activation through a PKC-dependent mechanism, whereas the late phase of bombesin-induced CCK secretion, that also requires PKC, appears to result from a MAPK-independent process.
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PMID:Bombesin stimulates cholecystokinin secretion through mitogen-activated protein-kinase-dependent and -independent mechanisms in the enteroendocrine STC-1 cell line. 951 70


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