Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Paper
  • Published:

Oxidation of a critical thiol residue of the adenine nucleotide translocator enforces Bcl-2-independent permeability transition pore opening and apoptosis

Abstract

Mitochondrial membrane permeabilization is a critical event in the process leading to physiological or chemotherapy-induced apoptosis. This permeabilization event is at least in part under the control of the permeability transition pore complex (PTPC), which interacts with oncoproteins from the Bcl-2 family as well as with tumor suppressor proteins from the Bax family, which inhibit or facilitate membrane permeabilization, respectively. Here we show that thiol crosslinking agents including diazenedicarboxylic acid bis 5N,N-dimethylamide (diamide), dithiodipyridine (DTDP), or bis-maleimido-hexane (BMH) can act on the adenine nucleotide translocator (ANT), one of the proteins within the PTPC. ANT alone reconstituted into artificial lipid bilayers suffices to confer a membrane permeabilization response to thiol crosslinking agents. Diamide, DTDP, and BMH but not tert-butylhydroperoxide or arsenite cause the oxidation of a critical cysteine residue (Cys 56) of ANT. Thiol modification within ANT is observed in intact cells, isolated mitochondria, and purified ANT. Recombinant Bcl-2 fails to prevent thiol modification of ANT. Concomitantly, a series of different thiol crosslinking agents (diamide, DTDP, and BMH, phenylarsine oxide) but not tert-butylhydroperoxide or arsenite induce mitochondrial membrane permeabilization and cell death irrespective of the expression level of Bcl-2. These data indicate that thiol crosslinkers cause a covalent modification of ANT which, beyond any control by Bcl-2, leads to mitochondrial membrane permeabilization and cell death.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7

Similar content being viewed by others

Abbreviations

ANT:

adenine nucleotide translocator

BMH:

bis-maleimido-hexane

CsA:

cyclosporin A

diamide:

diazenedicarboxylic acid bis 5N,N-dimethylamide

ΔΨm:

mitochondrial transmembrane potential

DiOC6(3):

3, 3′dihexyloxacarbocyanine iodide

dithiodipyridine:

DTDP

HE:

hydroethidine

4-MUP:

4-methylumbelliferylphosphate

4-MU:

4-methylumbelliferone

PI:

propidium iodide

PT:

permeability transition

PTPC:

permeability transition pore complex

PYM:

N-(1-pyrenil)maleimide

t-BHP:

tert-butylhydroperoxide

VDAC:

voltage dependent anion channel

References

  • Beutner G, Ruck A, Riede B and Brdiczka D . 1998 Biochim Biophys Acta - Biomembr 1368: 7–18.

  • Beyer K and Nuscher B . 1996 Biochemistry 35: 15784–15790.

  • Brenner C, Cadiou H, Vieira HLA, Zamzami N, Marzo I, Xie Z, Leber B, Andrews D, Duclohier H, Reed JC and Kroemer G . 1999 Oncogene, in press.

  • Brustovetsky N and Klingenberg M . 1996 Biochemistry 35: 8483–8488.

  • Costantini P, Chernyak BV, Petronilli V and Bernardi P . 1995a FEBS Lett 362: 239–242.

  • Costantini P, Chernyak BV, Petronilli V and Bernardi P . 1996 J Biol Chem 271: 6746–6751.

  • Costantini P, Colonna R and Bernardi P . 1998 Biochim Biophys Acta 1365: 385–392.

  • Costantini P, Petronilli V, Colonna R and Bernardi P . 1995b Toxicology 99: 77–88.

  • Crompton M, Virji S and Ward JM . 1998 Eur J Biochem 258: 729–735.

  • Giron-Calle J and Schmid HH . 1996 Biochemistry 35: 15440–15446.

  • Green DR and Reed JC . 1998 Science 281: 1309–1312.

  • Halestrap AP, Woodfield KY and Connern CP . 1997 J Biol Chem 272: 3346–3354.

  • Heiskanen KM, Bhat MB, Wang HW, Ma JJ and Nieminen AL . 1999 J Biol Chem 274: 5654–5658.

  • Hirsch T, Decaudin D, Susin SA, Marchetti P, Larochette N, Resche-Rigon M and Kroemer G . 1998 Exp Cell Res 241: 426–434.

  • Hockenbery DM, Oltvai ZN, Yin X-M, Milliman CL and Korsmeyer SJ . 1993 Cell 75: 241–251.

  • Kane DJ, Sarafian TA, Anton R, Hahn H, Gralla EB, Valentine JS, Örd T and Bredesen DE . 1993 Science 262: 1274–1277.

  • Klingenberg M . 1993 J Bioenerg Biomembr 25: 447–457.

  • Kluck RM, Bossy-Wetzel E, Green DR and Newmeyer DD . 1997 Science 275: 1132–1136.

  • Kroemer G, Dallaporta B and Resche-Rigon M . 1998 Annu Rev Physiol 60: 619–642.

  • Larochette N, Decaudin D, Jacotot E, Brenner C, Marzo I, Susin SA, Zamzami N, Xie Z, Reed JC and Kroemer G . 1999 Exp Cell Res 249: 413–421.

  • Liu XS, Kim CN, Yang J, Jemmerson R and Wang X . 1996 Cell 86: 147–157.

  • Majima E, Ikawa K, Takeda M, Hashimoto M, Shinohara Y and Terada H . 1995 J Biol Chem 270: 29548–29554.

  • Mancini M, Nicholson DW, Roy S, Thornberry NA, Peterson EP, Casciola-Rosen LA and Rosen A . 1998 J Cell Biol 140: 1485–1495.

  • Marchetti P, Castedo M, Susin SA, Zamzami N, Hirsch T, Haeffner A, Hirsch F, Geuskens M and Kroemer G . 1996a J Exp Med 184: 1155–1160.

  • Marchetti P, Hirsch T, Zamzami N, Castedo M, Decaudin D, Susin SA, Masse B and Kroemer G . 1996b J Immunol 157: 4830–4836.

  • Marzo I, Brenner C, Zamzami N, Jürgensmeier J, Susin SA, Vieira HLA, Prévost M-C, Xie Z, Mutsiyama S, Reed JC and Kroemer G . 1998a Science 281: 2027–2031.

  • Marzo I, Brenner C, Zamzami N, Susin SA, Beutner G, Brdiczka D, Rémy R, Xie Z-H, Reed JC and Kroemer G . 1998b J Exp Med 187: 1261–1271.

  • McEnery MW, Snowman AM, Trifiletti RR and Snyder SH . 1992 Proc Natl Acad Sci USA 89: 3170–3174.

  • Narita M, Shimizu S, Ito T, Chittenden T, Lutz RJ, Matsuda H and Tsujimoto Y . 1998 Proc Natl Acad Sci USA 95: 14681–14686.

  • Nicolli A, Basso E, Petronilli V, Wenger RM and Bernardi P . 1996 J Biol Chem 271: 2185–2192.

  • Palmieri F . 1994 FEBS Lett 346: 48–54.

  • Pedersen PL, Grennawalt JW, Reynafarje B, Hullihen J, Decker GL, Soper JW and Bustamente E . 1978 Meth Cell Biol 20: 411–481.

  • Ravagnan L, Marzo I, Costantini P, Susin SA, Zamzami N, Petit PX, Hirsch F, Poupon M-F, Miccoli L, Xie Z, Reed JC and Kroemer G . 1999 Oncogene 18: 2537–2546.

  • Rück A, Dolder M, Wallimann T and Brdiczka D . 1998 FEBS Lett 426: 97–101.

  • Shimizu S, Narita M and Tsujimoto Y . 1999 Nature 399: 483–487.

  • Susin SA, Larochette N, Geuskens M and Kroemer G . 1999a Meth Enzymol, in press.

  • Susin SA, Lorenzo HK, Zamzami N, Marzo I, Larochette N, Alzari PM and Kroemer G . 1999b J Exp Med 189: 381–394.

  • Susin SA, Lorenzo HK, Zamzami N, Marzo I, Snow BE, Brothers GM, Mangion J, Jacotot E, Costantini P, Loeffler M, Larochette N, Goodlett DR, Aebersold R, Siderovski DP, Penninger JM and Kroemer G . 1999c Nature 397: 441–446.

  • Susin SA, Zamzami N, Castedo M, Hirsch T, Marchetti P, Macho A, Daugas E, Geuskens M and Kroemer G . 1996 J Exp Med 184: 1331–1342.

  • vander Heiden MG, Chandal NS, Williamson EK, Schumacker PT and Thompson CB . 1997 Cell 91: 627–637.

  • Woodfield K, Ruck A, Brdiczka D and Halestrap AP . 1998 Biochem J 336: 287–290.

  • Yang J, Liu X, Bhalla K, Kim CN, Ibrado AM, Cai J, Peng T-I, Jones DP and Wang X . 1997 Science 275: 1129–1132.

  • Zamzami N, Marchetti P, Castedo M, Decaudin D, Macho A, Hirsch T, Susin SA, Petit PX, Mignotte B and Kroemer G . 1995 J Exp Med 182: 367–377.

  • Zamzami N, Marzo I, Susin SA, Brenner C, Larochette N, Marchetti P, Reed J, Kofler R and Kroemer G . 1998 Oncogene 16: 1055–1063.

  • Zamzami N, Susin SA, Marchetti P, Hirsch T, Gómez-Monterrey I, Castedo M and Kroemer G . 1996 J Exp Med 183: 1533–1544.

Download references

Acknowledgements

We thank Drs John Reed and Z Xue (The Burnham Institute, La Jolla, CA, USA) for the gift of recombinant Bcl-2 protein. This project has received a special grant from the Ligue National contre le Cancer (France). The work has also been supported by grants from ANRS, ARC, CNRS, FRM, French Ministry for Science, and Sidaction (to G Kroemer). P Costantini received a Fondation por la Recherche Medicale fellowship and HLA Vieira a fellowship from the Fundação para a Ciência e a Tecnolgia PRAXIS XXI, Portugal.

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Costantini, P., Belzacq, AS., Vieira, H. et al. Oxidation of a critical thiol residue of the adenine nucleotide translocator enforces Bcl-2-independent permeability transition pore opening and apoptosis. Oncogene 19, 307–314 (2000). https://doi.org/10.1038/sj.onc.1203299

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.onc.1203299

Keywords

This article is cited by

Search

Quick links