Abstract
Following the screening of a battery of distinct small-interfering RNAs that target various components of the apoptotic machinery, we found that knockdown of the voltage-dependent anion channel 1 (VDAC1) was particularly efficient in preventing cell death induced by cisplatin (CDDP) in non-small cell lung cancer cells. Both the downregulation of VDAC1 and its chemical inhibition with 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid reduced the apoptosis-associated modifications induced by CDDP, including mitochondrial transmembrane potential dissipation and plasma membrane permeabilization. VDAC1 inhibition strongly reduced the CDDP-induced conformational activation of Bax, yet had no discernible effect on the activation of Bak, suggesting that VDAC1 acts downstream of Bak and upstream of Bax. Accordingly, knockdown of Bak abolished the activation of Bax, whereas Bax downregulation had no effect on Bak activation. In VDAC1-depleted cells, the failure of CDDP to activate Bax could be reversed by means of the Bcl-2/Bcl-XL antagonist ABT-737, which concomitantly restored CDDP cytotoxicity. Altogether, these results delineate a novel pathway for the induction of mitochondrial membrane permeabilization (MMP) in the course of CDDP-induced cell death that involves a hierarchical contribution of Bak, VDAC1 and Bax. Moreover, our data suggest that VDAC1 may act as a facultative regulator/effector of MMP, depending on the initial cytotoxic event.
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Abbreviations
- ΔΨm:
-
mitochondrial transmembrane potential
- BH3:
-
Bcl-2 homology domain 3
- BRCA1:
-
breast cancer 1, early onset
- CDDP:
-
cisplatin
- Cyt c:
-
cytochrome c
- DAPI:
-
4′,6-diamidino-2-phenylindole dihydrochloride
- DIDS:
-
4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid
- DiOC6(3):
-
3,3′dihexiloxalocarbocyanine iodide
- ERCC1:
-
excision repair cross-complementing rodent repair deficiency, complementation group 1
- H2O2:
-
hydrogen peroxide
- HE:
-
hydroethidine
- IMS:
-
intermembrane space
- LDH:
-
lactate dehydrogenase
- MMP:
-
mitochondrial membrane permeabilization
- NAC:
-
N-acetylcysteine
- NSCLC:
-
non-small cell lung cancer
- OM:
-
mitochondrial outer membrane
- PARP-1:
-
poly (ADP-ribose) polymerase 1
- PI:
-
propidium iodide
- ROS:
-
reactive oxygen species
- siRNA:
-
small-interfering RNA
- tBHP:
-
tert-butylhydroperoxide
- VDAC1:
-
voltage-dependent anion channel 1
- WST-1:
-
4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate.
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Acknowledgements
This work has been supported by a special grant from Ligue National contre le cancer (équipe labellisée), as well as by grants from Agence Nationale de Recherche, Agence Nationale pour la Recherche sur le Sida, Cancéropôle Ile-de-France, Fondation pour la Recherche Médicale, Institut National du Cancer, European Commission (ApoSys, RIGHT, Active p53, Trans-Death, DeathTrain, ChemoRes) and Sidaction. NT is recipient of an FRM PhD fellowship. OK is recipient of an EMBO PhD fellowship. LS is funded by a DeathTrain PhD fellowship. EM is recipient of a DeathTrain PhD student fellowship.
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Tajeddine, N., Galluzzi, L., Kepp, O. et al. Hierarchical involvement of Bak, VDAC1 and Bax in cisplatin-induced cell death. Oncogene 27, 4221–4232 (2008). https://doi.org/10.1038/onc.2008.63
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DOI: https://doi.org/10.1038/onc.2008.63
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