Abstract
Granulocyte/macrophage colony-stimulating factor (GM-CSF) inhibits Fas-induced apoptosis of neutrophils. However, the exact step in the apoptotic pathway blocked by GM-CSF remained unclear. Here, we found that pretreatment of neutrophils with GM-CSF inhibits the recruitment of Fas-associated protein with death domain (FADD) to Fas, abolishing the formation of the death-inducing signaling complex required for Fas-induced apoptosis. Two-dimensional electrophoresis revealed that GM-CSF modifies the ratio of FADD subspecies. These GM-CSF-triggered changes were abrogated, and Fas-induced apoptosis was restored by an inhibitor of classical protein kinase C (PKC), Go6976, and by the combination of a phosphatidylinositol 3-kinase (PI-3K) inhibitor, LY294002, and an inhibitor of mitogen-activated protein kinase kinase (MEK)1, PD98059. Go6976 blocked GM-CSF-elicited phosphorylation of Akt/PKB and extracellular signal-regulated kinase (ERK)1/2. These results indicated that GM-CSF suppresses Fas-induced neutrophil apoptosis by inhibiting FADD binding to Fas, through redundant actions of PI-3K and MEK1-ERK1/2 pathways downstream of classical PKC.
MeSH terms
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Adaptor Proteins, Signal Transducing / drug effects
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Adaptor Proteins, Signal Transducing / metabolism*
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Adult
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Apoptosis / drug effects
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Apoptosis / immunology
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Apoptosis / physiology*
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Caspase 8
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Caspases / metabolism
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Cells, Cultured
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Down-Regulation / drug effects
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Down-Regulation / physiology
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Enzyme Inhibitors / pharmacology
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Fas-Associated Death Domain Protein
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Feedback, Physiological / drug effects
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Feedback, Physiological / physiology
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Granulocyte-Macrophage Colony-Stimulating Factor / pharmacology*
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Humans
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MAP Kinase Kinase 1 / antagonists & inhibitors
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MAP Kinase Kinase 1 / metabolism
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Mitogen-Activated Protein Kinase 1 / antagonists & inhibitors
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Mitogen-Activated Protein Kinase 1 / metabolism
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Mitogen-Activated Protein Kinase 3 / antagonists & inhibitors
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Mitogen-Activated Protein Kinase 3 / metabolism
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Neutrophils / drug effects
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Neutrophils / enzymology*
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Neutrophils / immunology
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Phosphatidylinositol 3-Kinases / drug effects
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Phosphatidylinositol 3-Kinases / metabolism
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Phosphorylation / drug effects
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Protein Binding / drug effects
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Protein Kinase C / antagonists & inhibitors
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Protein Kinase C / metabolism
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Protein Serine-Threonine Kinases / antagonists & inhibitors
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Protein Serine-Threonine Kinases / metabolism
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Proto-Oncogene Proteins / antagonists & inhibitors
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Proto-Oncogene Proteins / metabolism
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Proto-Oncogene Proteins c-akt
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Signal Transduction / drug effects
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Signal Transduction / immunology
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Signal Transduction / physiology*
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Time Factors
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fas Receptor / drug effects
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fas Receptor / metabolism*
Substances
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Adaptor Proteins, Signal Transducing
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Enzyme Inhibitors
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FADD protein, human
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Fas-Associated Death Domain Protein
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Proto-Oncogene Proteins
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fas Receptor
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Granulocyte-Macrophage Colony-Stimulating Factor
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AKT1 protein, human
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Protein Serine-Threonine Kinases
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Proto-Oncogene Proteins c-akt
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Protein Kinase C
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Mitogen-Activated Protein Kinase 1
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Mitogen-Activated Protein Kinase 3
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MAP Kinase Kinase 1
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MAP2K1 protein, human
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CASP8 protein, human
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Caspase 8
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Caspases