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Changes in intramitochondrial and cytosolic pH: early events that modulate caspase activation during apoptosis
S Matsuyama1, J Llopis, Q L Deveraux
1Programme on Apoptosis and Cell Death Regulation, The Burnham Institute, 10901 North Torrey Pines Road, La Jolla, California 92037, USA.
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.
Insights
Mitochondria-dependent apoptosis involves rapid pH changes, including mitochondrial alkalinization and cytosol acidification. This pH shift is crucial for cytochrome c release and caspase activation, regulating the apoptotic pathway.
Area of Science:
- Cell Biology
- Biochemistry
- Apoptosis Research
Background:
- Mitochondria are key regulators of apoptosis, releasing cytochrome c (cytC) to activate caspases.
- The precise role of intracellular pH dynamics in mitochondria-mediated apoptosis remains incompletely understood.
Purpose of the Study:
- To investigate the role of mitochondrial pH changes in the initiation and regulation of apoptosis.
- To determine if altered intracellular pH influences cytochrome c release and caspase activation.
Main Methods:
- Utilized pH-sensitive green fluorescent protein (GFP) to monitor mitochondrial and cytosolic pH.
- Applied various apoptotic stimuli (Bax, staurosporine, UV irradiation, Fas) and inhibitors (Bcl-2, oligomycin, caspase inhibitors).
- Conducted experiments in mammalian cells and yeast cells, including FoF1-ATPase/H+-pump-deficient strains.
Main Results:
- Mitochondria-dependent apoptotic stimuli induced rapid mitochondrial alkalinization and cytosol acidification, preceding cytochrome c release.
- These pH changes were inhibited by Bcl-2 and oligomycin, but not by caspase inhibitors.
- Cytosolic caspase activation was pH-dependent, with maximal activity at acidic pH.
Conclusions:
- Mitochondria-mediated alterations in intracellular pH are an early event in the apoptotic pathway.
- Cytosolic acidification appears to be a critical factor for efficient caspase activation.
- This study highlights the importance of the mitochondrial FoF1-ATPase/H+-pump in regulating apoptosis via pH modulation.