Abstract
The release of cytochrome c from mitochondria results in the formation of an Apaf-1-caspase-9 apoptosome and induces the apoptotic protease cascade by activation of procaspase-3. The present studies demonstrate that heat shock protein 90 (Hsp90) forms a cytosolic complex with Apaf-1 and thereby inhibits the formation of the active complex. Immunodepletion of Hsp90 depletes Apaf-1 and thereby inhibits cytochrome c-mediated activation of caspase-9. Addition of purified Apaf-1 to Hsp90-depleted cytosolic extracts restores cytochrome c-mediated activation of procaspase-9. We also show that Hsp90 inhibits cytochrome c-mediated oligomerization of Apaf-1 and thereby activation of procaspase-9. Furthermore, treatment of cells with diverse DNA-damaging agents dissociates the Hsp90-Apaf-1 complex and relieves the inhibition of procaspase-9 activation. These findings provide the first evidence for a negative cytosolic regulator of cytochrome c-dependent apoptosis and for involvement of a chaperone in the caspase cascade.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Amino Acid Sequence
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Apoptosis
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Apoptotic Protease-Activating Factor 1
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Binding, Competitive
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Caspase 3
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Caspase 9
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Caspases / metabolism*
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Cell Line
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Cell-Free System
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Chromatography, Affinity
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Cytochrome c Group / chemistry
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Cytochrome c Group / metabolism*
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Cytoplasm / metabolism
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Cytosol / metabolism
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Dose-Response Relationship, Drug
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Electrophoresis, Polyacrylamide Gel
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Enzyme Activation
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Enzyme Precursors / metabolism*
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Gene Expression Regulation, Enzymologic*
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HSP90 Heat-Shock Proteins / metabolism*
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Humans
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Immunoblotting
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Isoenzymes / metabolism
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Models, Biological
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Molecular Sequence Data
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Precipitin Tests
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Protein Binding
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Protein Kinase C / metabolism
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Protein Kinase C-delta
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Proteins / chemistry
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Proteins / metabolism*
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Sequence Homology, Amino Acid
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Transfection
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U937 Cells
Substances
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APAF1 protein, human
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Apoptotic Protease-Activating Factor 1
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Cytochrome c Group
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Enzyme Precursors
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HSP90 Heat-Shock Proteins
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Isoenzymes
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Proteins
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PRKCD protein, human
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Protein Kinase C
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Protein Kinase C-delta
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CASP3 protein, human
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CASP9 protein, human
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Caspase 3
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Caspase 9
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Caspases