Abstract
Many pro-apoptotic signals activate caspase-9, an initiator protease that activates caspase-3 and downstream caspases to initiate cellular destruction. However, survival signals can impinge on this pathway and suppress apoptosis. Activation of the Ras-Raf-MEK-ERK mitogen-activated protein kinase (MAPK) pathway is associated with protection of cells from apoptosis and inhibition of caspase-3 activation, although the targets are unknown. Here, we show that the ERK MAPK pathway inhibits caspase-9 activity by direct phosphorylation. In mammalian cell extracts, cytochrome c-induced activation of caspases-9 and -3 requires okadaic-acid-sensitive protein phosphatase activity. The opposing protein kinase activity is overcome by treatment with the broad-specificity kinase inhibitor staurosporine or with inhibitors of MEK1/2. Caspase-9 is phosphorylated at Thr 125, a conserved MAPK consensus site targeted by ERK2 in vitro, in a MEK-dependent manner in cells stimulated with epidermal growth factor (EGF) or 12-O-tetradecanoylphorbol-13-acetate (TPA). Phosphorylation at Thr 125 is sufficient to block caspase-9 processing and subsequent caspase-3 activation. We suggest that phosphorylation and inhibition of caspase-9 by ERK promotes cell survival during development and tissue homeostasis. This mechanism may also contribute to tumorigenesis when the ERK MAPK pathway is constitutively activated.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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3T3 Cells
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Animals
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Apoptosis / drug effects
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Apoptosis / physiology*
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Base Sequence / genetics
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Caspase 3
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Caspase 9
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Caspases / metabolism*
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Cell Survival / drug effects
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Cell Survival / physiology*
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Cell Transformation, Neoplastic / genetics
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Cell Transformation, Neoplastic / metabolism*
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Cytochrome c Group / drug effects
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Cytochrome c Group / metabolism
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Enzyme Inhibitors / pharmacology
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Epidermal Growth Factor / pharmacology
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Eukaryotic Cells / drug effects
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Eukaryotic Cells / enzymology*
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HeLa Cells
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Humans
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MAP Kinase Kinase 1
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Mice
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Mitogen-Activated Protein Kinase Kinases / metabolism
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Mitogen-Activated Protein Kinases / antagonists & inhibitors
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Mitogen-Activated Protein Kinases / metabolism*
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Molecular Sequence Data
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Phosphorylation / drug effects
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Protein Serine-Threonine Kinases / metabolism
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Pyridines / pharmacology
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Recombinant Fusion Proteins
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Signal Transduction / drug effects
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Signal Transduction / physiology
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Threonine / metabolism
Substances
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Cytochrome c Group
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Enzyme Inhibitors
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Pyridines
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Recombinant Fusion Proteins
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tris(2-pyridylmethyl)amine
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Threonine
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Epidermal Growth Factor
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Protein Serine-Threonine Kinases
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Mitogen-Activated Protein Kinases
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MAP Kinase Kinase 1
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MAP2K1 protein, human
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Map2k1 protein, mouse
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Mitogen-Activated Protein Kinase Kinases
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CASP3 protein, human
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CASP9 protein, human
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Casp3 protein, mouse
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Casp9 protein, mouse
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Caspase 3
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Caspase 9
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Caspases