Abstract
The pro-apoptotic Bcl-2 protein Bax can permeabilize the outer mitochondrial membrane and therefore commit human cells to apoptosis. Bax is regulated by constant translocation to the mitochondria and retrotranslocation back into the cytosol. Bax retrotranslocation depends on pro-survival Bcl-2 proteins and stabilizes inactive Bax. Here we show that Bax retrotranslocation shuttles membrane-associated and membrane-integral Bax from isolated mitochondria. We further discover the mitochondrial porin voltage-dependent anion channel 2 (VDAC2) as essential component and platform for Bax retrotranslocation. VDAC2 ensures mitochondria-specific membrane association of Bax and in the absence of VDAC2 Bax localizes towards other cell compartments. Bax retrotranslocation is also regulated by nucleotides and calcium ions, suggesting a potential role of the transport of these ions through VDAC2 in Bax retrotranslocation. Together, our results reveal the unanticipated bifunctional role of VDAC2 to target Bax specifically to the mitochondria and ensure Bax inhibition by retrotranslocation into the cytosol.
Highlights
Mitochondrial apoptosis is the most common form of programmed cell death[1] and involves proteins of the B-cell lymphoma-2 (Bcl-2) family
To extend the analysis of Bax retrotranslocation, we developed an in organelle retrotranslocation assay, monitoring the shuttling of Bax from purified mitochondria into the supernatant until translocation to the mitochondria and retrotranslocation from the mitochondria are in balance
Our results demonstrate that Bax retrotranslocation from the mitochondria into the cytosol depends on voltage-dependent anion channel 2 (VDAC2), indicating the possibility of Bax retrotranslocation induced by VDAC2 conformational changes[34,35,36] (Fig. 4C)
Summary
Mitochondrial apoptosis is the most common form of programmed cell death[1] and involves proteins of the B-cell lymphoma-2 (Bcl-2) family. Activation of the functionally redundant pro-apoptotic Bcl-2 proteins Bax and Bak leads to the permeabilization of the outer mitochondrial membrane (OMM) and subsequent release of intermembrane space (IMS) proteins into the cytoplasm[3]. Current evidence suggests that Bax, alone or in complex with other proteins, forms pores large enough to release IMS proteins[16]. Bax has been suggested to embed into the OMM, leading to the formation of lipidic pores explaining IMS protein release[18]. In the absence of VDAC2 Bax fails to target the OMM, suggesting a role of VDAC2 as Bax receptor in line with previous observations of a pro-apoptotic role of VDAC2 interactions with Bax/Bak. We conclude that VDAC2 has a bifunctional role to confer OMM-specific Bax recruitment and concomitant Bax inhibition via retrotranslocation
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