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BH3 death domain peptide induces cell type-selective mitochondrial outer membrane permeability
B M Polster1, K W Kinnally, G Fiskum
1Department of Anesthesiology, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
Abstract:
The BH3 domain is essential for the release of cytochrome c from mitochondria by pro-apoptotic Bcl-2 family proteins during apoptosis. This study tested the hypothesis that a Bax peptide that includes the BH3 domain can permeabilize the mitochondrial outer membrane and release cytochrome c in the absence of a permeability transition at the mitochondrial inner membrane. BH3 peptide (0.1-60 microm) released cytochrome c from mitochondria in the presence of physiological concentrations of ions in a cell type-selective manner, whereas a BH3 peptide with a single amino acid substitution was ineffective. The release of cytochrome c by BH3 peptide correlated with the presence of endogenous Bax at the mitochondria and its integral membrane insertion. Cytochrome c release was accompanied by adenylate kinase release, was not associated with mitochondrial swelling or substantial loss of electrical potential across the inner membrane, and was unaffected by inhibitors of the permeability transition pore. Cytochrome c release was, however, inhibited by Bcl-2. Although energy-coupled respiration was inhibited after the release of cytochrome c, mitochondria maintained membrane potential in the presence of ATP due to the reversal of the ATP synthase. Overall, results support the hypothesis that BH3 peptide releases cytochrome c by a Bax-dependent process that is independent of the mitochondrial permeability transition pore but regulated by Bcl-2.
Insights
A specific BH3 domain peptide triggers cytochrome c release from mitochondria, a key step in apoptosis. This Bax-dependent process bypasses the mitochondrial permeability transition pore and is regulated by Bcl-2.
Area of Science:
- Cell Biology
- Biochemistry
- Molecular Biology
Background:
- The BH3 domain of Bcl-2 family proteins is crucial for initiating apoptosis by releasing cytochrome c.
- Understanding the precise mechanism of cytochrome c release is vital for developing targeted therapies.
Purpose of the Study:
- To investigate if a Bax-derived BH3 peptide can induce mitochondrial outer membrane permeabilization and cytochrome c release.
- To determine if this release occurs independently of the mitochondrial permeability transition pore.
Main Methods:
- Treatment of isolated mitochondria with varying concentrations of BH3 peptide.
- Assessing cytochrome c and adenylate kinase release.
- Monitoring mitochondrial membrane potential and swelling.
- Evaluating the effect of Bcl-2 and permeability transition pore inhibitors.
Main Results:
- BH3 peptide induced cell-type-selective cytochrome c release, dependent on endogenous Bax.
- Cytochrome c release was independent of mitochondrial swelling and inner membrane potential loss.
- Bcl-2 inhibited BH3 peptide-induced cytochrome c release, while permeability transition pore inhibitors had no effect.
- Mitochondria maintained membrane potential post-cytochrome c release due to ATP synthase reversal.
Conclusions:
- BH3 peptide facilitates cytochrome c release via a Bax-dependent mechanism.
- This process is distinct from the mitochondrial permeability transition pore.
- Bcl-2 acts as a regulator, inhibiting this specific pathway of cytochrome c release.