Abstract
The heterologous expression of Bax, and other Bcl-2 family members, in the yeast Saccharomyces cerevisiae, has proved to be a valuable reporter system to investigate the molecular mechanisms underlying their interaction with mitochondria. By combining the co-expression of Bax and Bcl-xL mutants with analyzes of their localization and interaction in mitochondria and post-mitochondrial supernatants, we showed that the ability of Bax and Bcl-xL to interact is dependent both on Bax phosphorylation - mimicked by a substitution S184D - and by Bax and Bcl-xL localization. This, and previous data, provide the molecular basis for a model of dynamic equilibrium for Bax localization and activation, regulated both by phosphorylation and Bcl-xL.
Highlights
The pro-apoptotic protein Bax is at the core of the process of mitochondria-dependent apoptosis in mammals
By combining the co-expression of Bax and Bcl-xL mutants with analyzes of their localization and interaction in mitochondria and post-mitochondrial supernatants, we showed that the ability of Bax and Bcl-xL to interact is dependent both on Bax phosphorylation - mimicked by a substitution S184D - and by Bax and Bcl-xL localization
We previously reported that the phosphomimetic mutation S184D rendered Bax less stable in yeast, due to increased sensitivity to proteases: it was present at a lower level than BaxWT, but was restored at a normal level following the addition of the vacuolar protease inhibitor
Summary
The pro-apoptotic protein Bax is at the core of the process of mitochondria-dependent apoptosis in mammals. This widely accepted model still contains a number of gray areas, including the role of the very hydrophobic C-terminal helix α9, that was absent from the structural data of the Bax dimer [7], and of which the actual role in Bax interaction with mitochondria remains unclear: its absence does not prevent the mitochondrial localization of Bax, nor Bax-induced outer membrane permeabilization [14,15].
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