Abstract
Mitochondria trigger apoptosis by releasing caspase activators, including cytochrome c (cytC). Here we show, using a pH-sensitive green fluorescent protein (GFP), that mitochondria-dependent apoptotic stimuli (such as Bax, staurosporine and ultraviolet irradiation) induce rapid, Bcl-2-inhibitable mitochondrial alkalinization and cytosol acidification, followed by cytC release, caspase activation and mitochondrial swelling and depolarization. These events are not induced by mitochondria-independent apoptotic stimuli, such as Fas. Activation of cytosolic caspases by cytC in vitro is minimal at neutral pH, but maximal at acidic pH, indicating that mitochondria-induced acidification of the cytosol may be important for caspase activation; this finding is supported by results obtained from cells using protonophores. Cytosol acidification and cytC release are suppressed by oligomycin, a FoF1-ATPase/H +-pump inhibitor, but not by caspase inhibitors. Ectopic expression of Bax in wild-type, but not FoF1/H+-pump-deficient, yeast cells similarly results in mitochondrial matrix alkalinization, cytosol acidification and cell death. These findings indicate that mitochondria-mediated alteration of intracellular pH may be an early event that regulates caspase activation in the mitochondrial pathway for apoptosis.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Apoptosis* / drug effects
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Apoptosis* / radiation effects
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Carbonyl Cyanide m-Chlorophenyl Hydrazone / pharmacology
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Caspase Inhibitors
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Caspases / metabolism*
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Cell Line
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Cytochrome c Group / metabolism
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Cytosol / drug effects
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Cytosol / enzymology
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Cytosol / metabolism*
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Cytosol / radiation effects
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Deoxyadenine Nucleotides / pharmacology
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Enzyme Activation / drug effects
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Enzyme Activation / radiation effects
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Green Fluorescent Proteins
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Humans
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Luminescent Proteins / genetics
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Luminescent Proteins / metabolism
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Mitochondria / drug effects
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Mitochondria / enzymology
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Mitochondria / metabolism*
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Mitochondria / radiation effects
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Mitochondrial Swelling / drug effects
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Mitochondrial Swelling / radiation effects
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Mutation
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Oligomycins / pharmacology
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Proto-Oncogene Proteins / genetics
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Proto-Oncogene Proteins / metabolism
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Proto-Oncogene Proteins c-bcl-2 / genetics
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Proto-Oncogene Proteins c-bcl-2 / metabolism
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Proton-Translocating ATPases / antagonists & inhibitors
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Proton-Translocating ATPases / genetics
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Proton-Translocating ATPases / metabolism
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Saccharomyces cerevisiae / cytology
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Saccharomyces cerevisiae / enzymology
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / metabolism
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Staurosporine / antagonists & inhibitors
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Staurosporine / pharmacology
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Ultraviolet Rays
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bcl-2-Associated X Protein
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fas Receptor / physiology
Substances
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BAX protein, human
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Caspase Inhibitors
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Cytochrome c Group
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Deoxyadenine Nucleotides
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Luminescent Proteins
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Oligomycins
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Proto-Oncogene Proteins
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Proto-Oncogene Proteins c-bcl-2
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bcl-2-Associated X Protein
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fas Receptor
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Green Fluorescent Proteins
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Carbonyl Cyanide m-Chlorophenyl Hydrazone
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Caspases
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Proton-Translocating ATPases
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Staurosporine
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2'-deoxyadenosine triphosphate