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Query: UNIPROT:P04141 (granulocyte-macrophage colony-stimulating factor)
 6,790 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mast cell growth factor (MGF, the ligand for c-kit receptor) can stimulate proliferation of factor dependent myeloid cell line, M07e, and MGF synergizes with granulocyte-macrophage colony-stimulating factor (GM-CSF) or IL-3 in this effect. The effect of MGF on protein tyrosine kinase activity in M07e cells was investigated by immunoblotting with anti-phosphotyrosine mAb and this was compared with effects of GM-CSF. MGF stimulation rapidly induced or enhanced at least 12 tyrosine phosphorylated bands. Major bands had molecular weights of 145, 120, 110, 98, 62, 55 and 42 kD. P145, the most prominent phosphorylated protein, was identified as c-kit product using anti-c-kit-mAb (YB5.B8), suggesting ligand-dependent receptor autophosphorylation. Five of six tyrosine phosphorylated bands induced or enhanced by GM-CSF stimulation comigrated with those tyrosine phosphorylated by MGF (138, 120, 76, 55 and 42 kD). P42 was identified, at least in part, as mitogen-activated protein (MAP) kinase. MGF induced tyrosine phosphorylation of a complex of GTPase-activating protein (GAP, 120 kD) and GAP associated proteins (p62/p190) as detected by anti-GAP Ab immunoprecipitation followed by immunoblotting with anti-phosphotyrosine mAb. GM-CSF also stimulated slightly but consistently tyrosine phosphorylation of GAP and p190 but not p62. Both MGF and GM-CSF enhanced Raf-1 phosphorylation and increased Raf-1 associated kinase activity in vitro. Phosphoamino acid analysis revealed Raf-1 phosphorylation by these two growth factors occurred almost exclusively on serine residues. No tyrosine phosphorylation of Raf-1 protein was detected. These data suggest shared and unshared components of signaling pathways of both factors, which may be involved in cell proliferation.
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PMID:Comparative analysis of signaling pathways between mast cell growth factor (c-kit ligand) and granulocyte-macrophage colony-stimulating factor in a human factor-dependent myeloid cell line involves phosphorylation of Raf-1, GTPase-activating protein and mitogen-activated protein kinase. 172 91

The ability of the receptor for the hematopoietic cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) to function in non-hematopoietic cells is unknown. NIH3T3 fibroblasts were transfected with cDNAs encoding the alpha and beta subunit of the human GM-CSF receptor and a series of stable transformants were isolated that bound GM-CSF with either low (KD = 860 - > 1000 pM) or high affinity (KD = 20-80 pM). Low affinity receptors were not functional. However, the reconstituted high affinity receptors were found to be capable of activating a number of signal transduction pathways, including tyrosine kinase activity, phosphorylation of Raf-1, and the transient induction of c-fos and c-myc mRNAs. The activation of protein tyrosine phosphorylation by GM-CSF in NIH3T3 cells was rapid (< 1 min) and transient (peaking at 5-20 min) and resulted in the phosphorylation of proteins of estimated molecular weights of 42, 44, 52/53 and 58-60 kDa. Some of these proteins co-migrated with proteins from myeloid cells that were phosphorylated on tyrosine residues in response to GM-CSF. In particular, p42 and p44 were identified as mitogen-activated protein kinases (MAP kinases), and the phosphorylation on tyrosine residues of p42 and p44 MAP kinases occurred at the same time as the phosphorylation of Raf-1. However, despite evidence for activation of many mitogenic signal transduction molecules, GM-CSF did not induce significant proliferation of transfected NIH3T3 cells. These results suggest that murine fibroblasts contain signal transducing molecules that can effectively interact with the human GM-CSF receptor, and that are sufficient to activate at least some of the same signal transduction pathways this receptor activates in myeloid cells, including activation of one or more tyrosine kinase(s). However, the level of activation of signal transduction is either below a threshold of necessary activity or at least one mitogenic signal necessary for proliferation is missing.
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PMID:The human granulocyte-macrophage colony-stimulating factor receptor is capable of initiating signal transduction in NIH3T3 cells. 768 77

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

Transfected Jurkat cells overexpressing extracellular signal-regulated kinase (ERK1), also referred to as mitogen-activated protein (MAP) kinase, were selected by Western blotting assay using anti-ERK1 and antiphosphotyrosine antibodies in combination with a functional MAP kinase assay. We then asked whether enhanced ERK1 expression had any effect on induction of T-cell cytokine genes. The results show that overexpression of ERK1 enhances expression of T-cell interleukin-2 (IL-2), IL-3, and granulocyte-macrophage colony-stimulating factor mRNA; no change was seen in expression of the alpha-actin gene. DNA-binding activities of the transcription factors AP1, NF-AT, and NF-kB were specifically increased twofold to fourfold in ERK1-overexpressing clones relative to nontransformed or vector-transformed cells, whereas no enhancement of CK1-CK2 protein DNA binding activity was detected after ERK1 overexpression. Additionally, increased NF-AT DNA binding activity was associated with functional enhancement of NF-AT transactivating activity in ERK1-overexpressing cells. These results provide direct evidence for the role of MAP kinase in the regulation of cytokine gene expression and indicate that such regulation is likely mediated through the enhanced DNA binding activity of specific nuclear transcription factors.
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PMID:Overexpression of mitogen-activated protein kinase (ERK1) enhances T-cell cytokine gene expression: role of AP1, NF-AT, and NF-KB. 840 Feb 95

The survival and proliferation of the UT-7 human leukemic cell line is strictly dependent on the presence of either interleukin 3, granulocyte-macrophage colony-stimulating factor or erythropoietin. In these cells, erythropoietin stimulation led to the rapid phosphorylation of several proteins including the erythropoietin receptor and proteins with molecular masses around 45 kDa which could be mitogen-activated protein (MAP) kinases. Separation of cytosol from resting or erythropoietin-stimulated UT-7 cells by anion-exchange chromatography revealed two peaks of myelin basic protein kinase activity. The kinase activity of the first peak was independent of erythropoietin treatment of the cells and corresponded to an unidentified 50-kDa kinase, whereas the second peak was only present in erythropoietin-stimulated cells and corresponded to three forms of MAP kinases with molecular masses of 45, 44 and 42 kDa. The three forms were separated by hydrophobic chromatography and were shown to be activated in erythropoietin-stimulated cells. The 44-kDa and 42-kDa forms corresponded to extracellular signal-regulated kinase (ERK)-1 and ERK-2, respectively. Evidence was obtained showing that the 45-kDa form is not a shifted form of ERK-1 but corresponded to a less well defined form of MAP kinase which may be the previously described ERK-4. MAP kinase activation was detected after 1 min erythropoietin stimulation and remained detectable after more than 1 hour. A role for MAP kinase activation in erythropoietin-stimulated cell proliferation was suggested by the simultaneous inhibition of erythropoietin-induced MAP kinase stimulation and cell proliferation. The potential activator of MAP kinase, RAF-1, was hyperphosphorylated in erythropoietin-stimulated cells and its autophosphorylation activity was strongly increased. The protein adaptor Shc was heavily phosphorylated in UT-7 erythropoietin-stimulated cells and associated strongly with a unidentified 145-kDa protein. However, Shc bound poorly to the activated erythropoietin receptor and most Shc proteins were cytosolic in both unstimulated and erythropoietin-stimulated cells. In contrast, Grb2 associated efficiently with the activated erythropoietin receptor and a significant part of Grb2 was associated to a particulate subcellular fraction upon erythropoietin stimulation.
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PMID:The signal transduction pathway of erythropoietin involves three forms of mitogen-activated protein (MAP) kinase in UT7 erythroleukemia cells. 852 71

Hemopoietic cells respond to cytokines by initiating tyrosine phosphorylation of receptors and receptor-associated proteins, leading to the activation of numerous cytosolic and membrane associated enzymes, including phosphatidylinositol 3-OH kinase (PI 3-kinase). Recent reports have suggested that PI 3-kinase may serve as an upstream activator of mitogen-activated protein (MAP) kinase. After stimulation with interleukin-3 and granulocyte-macrophage colony-stimulating factor, we show here that inhibition of MAP kinase activity by two inhibitors of PI 3-kinase, wortmannin and LY-294002, does not correlate with their ability to inhibit PI 3-kinase or p70 S6 kinase phosphorylation. Complete inhibition of phosphatidylinositol 3,4,5-trisphosphate production occurred at approximately 100 nM WM or 25 microM LY-294002, but at these concentrations, WM significantly inhibited MAP kinase activation, while LY-294002 had virtually no effect on MAP kinase activity. Furthermore, WM does not inhibit phorbol ester-mediated MAP kinase activation, but LY-294002 does. Together these results suggest WM and LY-294002 are differentially inhibiting enzymes other than PI 3-kinase that function upstream of MAP kinase.
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PMID:Phosphatidylinositol 3-OH kinase activity is not required for activation of mitogen-activated protein kinase by cytokines. 866 37

The presence of a novel 38 kDa protein that is tyrosine phosphorylated in human neutrophils, a terminally differentiated cell, upon stimulation of these cells with low concentrations of lipopolysaccharide (LPS) in combination with serum has been demonstrated. This 38 kDa protein was identified as the mammalian homologue of HOG1 in yeast, the p38 mitogen-activated protein (MAP) kinase. This conclusion is based on the experimental findings that anti-phosphotyrosine (anti-PY) antibody immunoprecipitates a 38 kDa protein that is recognized by anti-p38 MAP kinase antibody, and conversely, anti-p38 MAP kinase antibody immunoprecipitates a 38 kDa protein that can be recognized by anti-PY antibody. Moreover, this tyrosine phosphorylated protein is found associated entirely with the cytosol. It was also found that this p38 MAP kinase is activated following stimulation of these cells with low concentrations of LPS in combination with serum. This conclusion is based on three experimental findings. First, soluble fractions isolated from LPS-stimulated cells phosphorylate heat shock protein 27 (hsp27) in an in vitro assay, and this effect is not inhibited by protein kinase C and protein kinase A inhibitor peptides. This effect is similar to the effect produced by the commercially available phosphorylated and activated MAPKAP kinase-2 (MAP kinase activated protein kinase-2). Secondly, a 27 kDa protein that aligns with a protein recognized by anti-hsp27 antibody is phosphorylated upon LPS stimulation of intact human neutrophils prelabelled with radioactive phosphate. Lastly, immune complex protein kinase assays, using [gamma-32P]ATP and activating transcription factor 2 (ATF2) as substrates, showed increased p38 MAP kinase activity from LPS-stimulated human neutrophils. The phosphorylation and activation of this p38 MAP kinase can be affected by both G-protein-coupled receptors such as platelet-activating factor (PAF) and non-G-protein-coupled receptors such as the cytokine-coupled receptors for granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumour necrosis factor alpha (TNF-alpha). The effect of low concentrations of PAF is greatly increased in cells pretreated with LPS. The tyrosine phosphorylation of the p38 MAP kinase is not restricted to stimuli that mediate their actions through membrane-associated receptors, but it can be affected by agents that bypass membrane-associated receptors such as the protein translation blocker anisomycin. While anisomycin is known to increase the tyrosine phosphorylation of the 54 kDa SAPK (stress-activated protein kinase), this is the first report that shows that anisomycin also tyrosine phosphorylates the p38 MAP kinase. Cytokine receptors that increase the tyrosine phosphorylation and activation of the erk1 and erk2 MAP kinases have less effect on this p38 MAP kinase than those that do not affect the erk1 and erk2 MAP kinases. The possible role of the p38 MAP kinase in the phosphorylation of cytosolic phospholipase A2 is discussed.
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PMID:Tyrosine phosphorylation and activation of a new mitogen-activated protein (MAP)-kinase cascade in human neutrophils stimulated with various agonists. 876 79

Activation and recruitment of eosinophils in allergic inflammation is in part mediated by chemoattractants and T-helper 2 (Th2)-derived cytokines. However, little is known concerning the signal transduction mechanisms by which this activation occurs. We have investigated tyrosine kinase-mediated activation of phosphatidylinositol 3-kinase (PI3K) and compared this with the activation of the p21ras-ERK signaling pathway in human eosinophils. The related cytokines interleukin-3 (IL-3), IL-5, and granulocyte-macrophage colony-stimulating factor (GM-CSF), all induced PI3K activity detected in antiphosphotyrosine immunoprecipitates. Furthermore, the chemoattractants platelet-activating factor (PAF), RANTES, and C5a were also able to induce phosphotyrosine-associated PI3K activity. Protein kinase B (PKB) is a downstream target of PI3K activation by growth factors. Induction of PKB phosphorylation in human eosinophils was transiently induced on activation with the cytokines IL-4 and IL-5, as well as the chemoattractants PAF, C5a, and RANTES showing a broad activation profile. Surprisingly, analysis of the activation of the mitogen-activated protein (MAP) kinases p44(ERK1) and p42(ERK2), showed that ERK2, but not ERK1, was transiently activated in human eosinophils after stimulation with IL-5 or PAF. Activation kinetics correlated with activation of p21ras by both cytokines and chemoattractants as measured by a novel assay for guanosine triphosphate (GTP)-loading. Finally, using specific inhibitors of both the p21ras-ERK and PI3K signaling pathways, a role was demonstrated for PI3K, but not p21ras-ERK, in activation of the serum-treated zymosan (STZ)-mediated respiratory burst in IL-5 and PAF-primed eosinophils. In summary, these data show that in human eosinophils, Th2-derived cytokines differentially activate both PI3K and MAP kinase signal transduction pathways with distinct functional consequences showing complex regulation of eosinophil effector functions.
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PMID:Analysis of signal transduction pathways in human eosinophils activated by chemoattractants and the T-helper 2-derived cytokines interleukin-4 and interleukin-5. 951 56

Incubation of human neutrophils with a chemotactic peptide [N-formylmethionyl-leucylphenylalanine (fMLP)] gave rise to an increase in the phosphoinositide 3-kinase (PI3K) activity, phosphorylation of p47phox and superoxide-anion (O2(-)) generation in the same fMLP-concentration-dependent manner. These responses to fMLP were markedly enhanced when the cells had been incubated for 10 min before the addition of fMLP with increasing concentrations of granulocyte-macrophage colony-stimulating factor (GM-CSF) that were only slightly effective themselves. Wortmannin, an inhibitor of PI3K, suppressed all of these fMLP actions in the same concentration-dependent manner in either GM-CSF-primed or non-primed cells. Sustained activation of protein kinase C by the addition of PMA caused marked phosphorylation of p47phox and respiratory burst itself without activation of PI3K. This strong action of PMA was not primed by GM-CSF. The chemotactic peptide was without effect in pertussis-toxin-treated cells, indicating that its actions are mediated by betagamma-subunits liberated from toxin-susceptible heterotrimeric Gi proteins (Gbetagamma). Thus one of the mechanisms of GM-CSF-mediated priming of fMLP-induced respiratory burst is synergistic activation of wortmannin-sensitive PI3K by Gbetagamma in the presence of tyrosine-phosphorylated proteins in GM-CSF-treated cells, as recently indicated in a cell-free system [Kurosu, Maehama, Okada, Yamamoto, Hoshino, Fukui, Ui, Hazeki and Katada (1997) J. Biol. Chem. 272, 24252-24256]. GM-CSF primed fMLP-induced MAP (mitogen-activated protein) kinase activation enormously as well. The MAP kinase activation was primed even in the presence of wortmannin, indicating that PI3K was not the sole site where tyrosine kinase-related and Gbetagamma-mediated intracellular signals converge to elicit the priming. The GM-CSF priming of fMLP-induced PI3K activation and O2(-) generation was much smaller in magnitude in neutrophils in which cAMP accumulated upon incubation with prostaglandin E1 than in the cells without the nucleotide accumulation. Thus the GM-CSF priming site, in addition to PI3K, might be just the target of cAMP-dependent protein kinase A in fMLP-initiated signalling cascades or could be localized immediately downstream thereof.
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PMID:Enhancement of chemotactic peptide-induced activation of phosphoinositide 3-kinase by granulocyte-macrophage colony-stimulating factor and its relation to the cytokine-mediated priming of neutrophil superoxide-anion production. 988 16

The role of granulocyte colony-stimulating factor (G-CSF) on neutrophilic differentiation of Me2SO-treated HL-60 cells was studied. G-CSF augmented the functional maturation of Me2SO-treated HL-60 cells in terms of both O-2-generating ability and expression of the formyl-methionyl-leucyl-phenylalanine receptor. G-CSF induced enhancement of cell growth in Me2SO-treated HL-60 cells. These results indicate that G-CSF is a potent enhancer for the differentiation and proliferation of Me2SO-treated HL-60 cells. G-CSF caused the activation of p70 S6 kinase but not mitogen-activated protein (MAP) kinase. On the other hand, G-CSF rapidly induced tyrosine phosphorylation of signal transducers and activators of transcription-3 (STAT3), but did not induce serine727 phosphorylation. From the analysis of confocal laser scanning fluorescence microscopy and differential centrifugation, it was clearly demonstrated that G-CSF induced nuclear translocation of tyrosine-phosphorylated STAT3. The G-CSF-dependent enhancement of neutrophilic differentiation in Me2SO-HL-60 cells was reversely inhibited by granulocyte-macrophage colony-stimulating factor (GM-CSF). Notably, in the presence of GM-CSF, G-CSF induced the tyrosine phosphorylation of STAT3 but failed to induce the nuclear translocation of tyrosine-phosphorylated STAT3. GM-CSF induced activation of not only p70 S6 kinase, but also of MAP kinase. Furthermore, GM-CSF caused the rapid serine727 phosphorylation of STAT3, both in the presence and absence of G-CSF. PD98059, an MEK1 inhibitor, inhibited the G-CSF-dependent serine727 phosphorylation of STAT3 and blocked the inhibitory effect of GM-CSF on G-CSF-dependent nuclear translocation of STAT3. These results suggest that G-CSF-dependent nuclear translocation of STAT3 coordinates with the promotion of neutrophilic differentiation in Me2SO-treated HL-60 cells.
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PMID:The role of STAT3 in granulocyte colony-stimulating factor-induced enhancement of neutrophilic differentiation of Me2SO-treated HL-60 cells. GM-CSF inhibits the nuclear translocation of tyrosine-phosphorylated STAT3. 1033 53


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