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.

  • Article
  • Published:

The apoptosis inhibitor Bcl-xL controls breast cancer cell migration through mitochondria-dependent reactive oxygen species production

Oncogene volume 39, pages 3056–3074 (2020)Cite this article

Subjects

Abstract

The Bcl-xL apoptosis inhibitor plays a major role in vertebrate development. In addition to its effect on apoptosis, Bcl-xL is also involved in cell migration and mitochondrial metabolism. These effects may favour the onset and dissemination of metastasis. However, the underlying molecular mechanisms remain to be fully understood. Here we focus on the control of cell migration by Bcl-xL in the context of breast cancer cells. We show that Bcl-xL silencing led to migration defects in Hs578T and MDA-MB231 cells. These defects were rescued by re-expressing mitochondria-addressed, but not endoplasmic reticulum-addressed, Bcl-xL. The use of BH3 mimetics, such as ABT-737 and WEHI-539 confirmed that the effect of Bcl-xL on migration did not depend on interactions with BH3-containing death accelerators such as Bax or BH3-only proteins. In contrast, the use of a BH4 peptide that disrupts the Bcl-xL/VDAC1 complex supports that Bcl-xL by acting on VDAC1 permeability contributes to cell migration through the promotion of reactive oxygen species production by the electron transport chain. Collectively our data highlight the key role of Bcl-xL at the interface between cell metabolism, cell death, and cell migration, thus exposing the VDAC1/Bcl-xL interaction as a promising target for anti-tumour therapy in the context of metastatic breast cancer.

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

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

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

Fig. 1: Bcl-xL protein levels in human breast epithelial cell lines.
Fig. 2: High bclx gene expression level is of poor prognosis in oestrogen receptor-positive patients.
Fig. 3: Bclx silencing impairs migration of mammary cancer cell lines.
Fig. 4: Bclx silencing inhibits the ability of 4T1cells to metastasize to the lung.
Fig. 5: Bcl-xL controls cell migration independently of its anti-apoptotic activity.
Fig. 6: Mitochondrial-, but not ER-targeted Bcl-xL, rescues cell migration.
Fig. 7: Mito-Bcl-xL forsters cell migration contrary to ER-Bcl-xL.
Fig. 8: Bcl-xL controls cell migration via its BH4 domain.
Fig. 9: Bclx silencing alters mitochondrial ATP generation.
Fig. 10: Cell motility alterations and drop in ROS production are closely correlated.
Fig. 11: Bclx silencing compromises calcium homeostasis.

Similar content being viewed by others

References

  1. Youle RJ, Strasser A. The BCL-2 protein family: opposing activities that mediate cell death. Nat Rev Mol Cell Biol. 2008;9:47–59.

    Article  CAS  PubMed  Google Scholar 

  2. Lopez J, Tait SWG. Mitochondrial apoptosis: killing cancer using the enemy within. Br J Cancer. 2015;112:957–62.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Bonneau B, Prudent J, Popgeorgiev N, Gillet G. Non-apoptotic roles of Bcl-2 family: the calcium connection. Biochim Biophys Acta. 2013;1833:1755–65.

    Article  CAS  PubMed  Google Scholar 

  4. Prudent J, Popgeorgiev N, Bonneau B, Thibaut J, Gadet R, Lopez J, et al. Bcl-wav and the mitochondrial calcium uniporter drive gastrula morphogenesis in zebrafish. Nat Commun. 2013;4:2330.

    Article  PubMed  CAS  Google Scholar 

  5. Li C, Wang X, Vais H, Thompson CB, Foskett JK, White C. Apoptosis regulation by Bcl-x(L) modulation of mammalian inositol 1,4,5-trisphosphate receptor channel isoform gating. Proc Natl Acad Sci USA. 2007;104:12565–70.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. White C, Li C, Yang J, Petrenko NB, Madesh M, Thompson CB, et al. The endoplasmic reticulum gateway to apoptosis by Bcl-X(L) modulation of the InsP3R. Nat Cell Biol. 2005;7:1021–8.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Huang H, Hu X, Eno CO, Zhao G, Li C, White C. An interaction between Bcl-xL and the voltage-dependent anion channel (VDAC) promotes mitochondrial Ca2+ uptake. J Biol Chem. 2013;288:19870–81.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Vander Heiden MG, Li XX, Gottleib E, Hill RB, Thompson CB, Colombini M. Bcl-xL promotes the open configuration of the voltage-dependent anion channel and metabolite passage through the outer mitochondrial membrane. J Biol Chem. 2001;276:19414–9.

    Article  Google Scholar 

  9. Vervliet T, Clerix E, Seitaj B, Ivanova H, Monaco G, Bultynck G. Modulation of Ca2+ signaling by anti-apoptotic B-cell lymphoma 2 proteins at the endoplasmic reticulum-mitochondrial interface. Front Oncol. 2017;7:75.

    Article  PubMed  PubMed Central  Google Scholar 

  10. Karczmarek-Borowska B, Filip A, Wojcierowski J, Smoleń A, Korobowicz E, Korszen-Pilecka I, et al. Estimation of prognostic value of Bcl-xL gene expression in non-small cell lung cancer. Lung Cancer. 2006;51:61–9.

    Article  PubMed  Google Scholar 

  11. Watanabe J, Kushihata F, Honda K, Mominoki K, Matsuda S, Kobayashi N. Bcl-xL overexpression in human hepatocellular carcinoma. Int J Oncol. 2002;21:515–9.

    CAS  PubMed  Google Scholar 

  12. Olopade OI, Adeyanju MO, Safa AR, Hagos F, Mick R, Thompson CB, et al. Overexpression of BCL-x protein in primary breast cancer is associated with high tumor grade and nodal metastases. Cancer J Sci Am. 1997;3:230–7.

    CAS  PubMed  Google Scholar 

  13. Martin SS, Ridgeway AG, Pinkas J, Lu Y, Reginato MJ, Koh EY, et al. A cytoskeleton-based functional genetic screen identifies Bcl-xL as an enhancer of metastasis, but not primary tumor growth. Oncogene. 2004;23:4641–5.

    Article  CAS  PubMed  Google Scholar 

  14. Fernández Y, España L, Mañas S, Fabra A, Sierra A. Bcl-xL promotes metastasis of breast cancer cells by induction of cytokines resistance. Cell Death Differ. 2000;7:350–9.

    Article  PubMed  CAS  Google Scholar 

  15. Keitel U, Scheel A, Thomale J, Halpape R, Kaulfuß S, Scheel C, et al. Bcl-xL mediates therapeutic resistance of a mesenchymal breast cancer cell subpopulation. Oncotarget. 2014;5:11778–91.

  16. Hager JH, Ulanet DB, Hennighausen L, Hanahan D. Genetic ablation of Bcl-x attenuates invasiveness without affecting apoptosis or tumor growth in a mouse model of pancreatic neuroendocrine cancer. PloS ONE. 2009;4:e4455.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  17. Du Y-CN, Lewis BC, Hanahan D, Varmus H. Assessing tumor progression factors by somatic gene transfer into a mouse model: Bcl-xL promotes islet tumor cell invasion. PLoS Biol. 2007;5:e276.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  18. Hwang K-T, Woo JW, Shin HC, Kim HS, Ahn SK, Moon H-G, et al. Prognostic influence of BCL2 expression in breast cancer. Int J Cancer J Int Cancer. 2012;131:E1109–1119.

    Article  CAS  Google Scholar 

  19. Neve RM, Chin K, Fridlyand J, Yeh J, Baehner FL, Fevr T, et al. A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes. Cancer Cell. 2006;10:515–27.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Schmitt E, Beauchemin M, Bertrand R. Nuclear colocalization and interaction between bcl-xL and cdk1(cdc2) during G2/M cell-cycle checkpoint. Oncogene. 2007;26:5851–65.

    Article  CAS  PubMed  Google Scholar 

  21. Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F. Genome engineering using the CRISPR-Cas9 system. Nat Protoc. 2013;8:2281–308.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Teng Y, Xie X, Walker S, White DT, Mumm JS, Cowell JK. Evaluating human cancer cell metastasis in zebrafish. BMC Cancer. 2013;13:453.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  23. Oltersdorf T, Elmore SW, Shoemaker AR, Armstrong RC, Augeri DJ, Belli BA, et al. An inhibitor of Bcl-2 family proteins induces regression of solid tumours. Nature. 2005;435:677–81.

    Article  CAS  PubMed  Google Scholar 

  24. Lessene G, Czabotar PE, Sleebs BE, Zobel K, Lowes KN, Adams JM, et al. Structure-guided design of a selective BCL-X(L) inhibitor. Nat Chem Biol. 2013;9:390–7.

    Article  CAS  PubMed  Google Scholar 

  25. Eno CO, Eckenrode EF, Olberding KE, Zhao G, White C, Li C. Distinct roles of mitochondria- and ER-localized Bcl-xL in apoptosis resistance and Ca2+ homeostasis. Mol Biol Cell. 2012;23:2605–18.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Bonneau B, Nougarède A, Prudent J, Popgeorgiev N, Peyriéras N, Rimokh R, et al. The Bcl-2 homolog Nrz inhibits binding of IP3 to its receptor to control calcium signaling during zebrafish epiboly. Sci Signal. 2014;7:ra14.

    Article  PubMed  CAS  Google Scholar 

  27. Banaszynski LA, Chen L, Maynard-Smith LA, Ooi AGL, Wandless TJ. A rapid, reversible, and tunable method to regulate protein function in living cells using synthetic small molecules. Cell. 2006;126:995–1004.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Monaco G, Decrock E, Arbel N, van Vliet AR, La Rovere R, De Smedt H, et al. The BH4 domain of anti-apoptotic Bcl-XL, but not that of the related Bcl-2, limits the voltage-dependent anion channel 1 (VDAC1)-mediated transfer of pro-apoptotic Ca2+ signals to mitochondria. J Biol Chem. 2015;290:9150–61.

  29. Shimizu S, Konishi A, Kodama T, Tsujimoto Y. BH4 domain of antiapoptotic Bcl-2 family members closes voltage-dependent anion channel and inhibits apoptotic mitochondrial changes and cell death. Proc Natl Acad Sci USA. 2000;97:3100–5.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Tsujimoto Y, Shimizu S. VDAC regulation by the Bcl-2 family of proteins. Cell Death Differ. 2000;7:1174–81.

    Article  CAS  PubMed  Google Scholar 

  31. Dogterom M, Koenderink GH. Actin-microtubule crosstalk in cell biology. Nat Rev Mol Cell Biol. 2019;20:38–54.

    Article  CAS  PubMed  Google Scholar 

  32. Senju Y, Lappalainen P. Regulation of actin dynamics by PI(4,5)P2 in cell migration and endocytosis. Curr Opin Cell Biol. 2019;56:7–13.

    Article  CAS  PubMed  Google Scholar 

  33. Angelova PR, Abramov AY. Functional role of mitochondrial reactive oxygen species in physiology. Free Radic Biol Med. 2016;100:81–5.

    Article  CAS  PubMed  Google Scholar 

  34. Kastl L, Sauer SW, Ruppert T, Beissbarth T, Becker MS, Süss D, et al. TNF-α mediates mitochondrial uncoupling and enhances ROS-dependent cell migration via NF-κB activation in liver cells. FEBS Lett. 2014;588:175–83.

    Article  CAS  PubMed  Google Scholar 

  35. Lim S-K, Choi YW, Lim IK, Park TJ. BTG2 suppresses cancer cell migration through inhibition of Src-FAK signaling by downregulation of reactive oxygen species generation in mitochondria. Clin Exp Metastasis. 2012;29:901–13.

    Article  CAS  PubMed  Google Scholar 

  36. Ludin A, Gur-Cohen S, Golan K, Kaufmann KB, Itkin T, Medaglia C, et al. Reactive oxygen species regulate hematopoietic stem cell self-renewal, migration and development, as well as their bone marrow microenvironment. Antioxid Redox Signal. 2014;21:1605–19.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Vander Heiden MG, Chandel NS, Schumacker PT, Thompson CB. Bcl-xL prevents cell death following growth factor withdrawal by facilitating mitochondrial ATP/ADP exchange. Mol Cell. 1999;3:159–67.

    Article  Google Scholar 

  38. Chen Y, Aon MA, Hsu Y-T, Soane L, Teng X, McCaffery JM, et al. Bcl-xL regulates mitochondrial energetics by stabilizing the inner membrane potential. J Cell Biol. 2011;195:263–76.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Hardwick JM, Chen Y, Jonas EA. Multipolar functions of BCL-2 proteins link energetics to apoptosis. Trends Cell Biol. 2012;22:318–28.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Prevarskaya N, Skryma R, Shuba Y. Calcium in tumour metastasis: new roles for known actors. Nat Rev Cancer. 2011;11:609–18.

    Article  CAS  PubMed  Google Scholar 

  41. Paupe V, Prudent J. New insights into the role of mitochondrial calcium homeostasis in cell migration. Biochem Biophys Res Commun. 2018;500:75–86.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Nguyen M-HT, Jafri MS. Mitochondrial calcium signaling and energy metabolism. Ann N Y Acad Sci. 2005;1047:127–37.

    Article  CAS  PubMed  Google Scholar 

  43. Griffiths EJ, Rutter GA. Mitochondrial calcium as a key regulator of mitochondrial ATP production in mammalian cells. Biochim Biophys Acta. 2009;1787:1324–33.

    Article  CAS  PubMed  Google Scholar 

  44. Ben-Hail D, Shoshan-Barmatz V. VDAC1-interacting anion transport inhibitors inhibit VDAC1 oligomerization and apoptosis. Biochim Biophys Acta. 2016;1863:1612–23.

    Article  CAS  PubMed  Google Scholar 

  45. Choi S, Chen Z, Tang LH, Fang Y, Shin SJ, Panarelli NC, et al. Bcl-xL promotes metastasis independent of its anti-apoptotic activity. Nat Commun. 2016;7:10384.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Vogler M, Hamali HA, Sun X-M, Bampton ETW, Dinsdale D, Snowden RT, et al. BCL2/BCL-X(L) inhibition induces apoptosis, disrupts cellular calcium homeostasis, and prevents platelet activation. Blood. 2011;117:7145–54.

    Article  CAS  PubMed  Google Scholar 

  47. González-García M, Pérez-Ballestero R, Ding L, Duan L, Boise LH, Thompson CB, et al. bcl-XL is the major bcl-x mRNA form expressed during murine development and its product localizes to mitochondria. Dev Camb Engl. 1994;120:3033–42.

    Google Scholar 

  48. Motoyama N, Wang F, Roth KA, Sawa H, Nakayama K, Nakayama K, et al. Massive cell death of immature hematopoietic cells and neurons in Bcl-x-deficient mice. Science. 1995;267:1506–10.

    Article  CAS  PubMed  Google Scholar 

  49. Johnson BW, Cepero E, Boise LH. Bcl-xL inhibits cytochrome c release but not mitochondrial depolarization during the activation of multiple death pathways by tumor necrosis factor-alpha. J Biol Chem. 2000;275:31546–53.

    Article  CAS  PubMed  Google Scholar 

  50. Popgeorgiev N, Jabbour L, Gillet G. Subcellular localization and dynamics of the Bcl-2 family of proteins. Front Cell Dev Biol. 2018;6:13.

    Article  PubMed  PubMed Central  Google Scholar 

  51. Yang J, Vais H, Gu W, Foskett JK Biphasic regulation of InsP3 receptor gating by dual Ca2+ release channel BH3-like domains mediates Bcl-xL control of cell viability. Proc Natl Acad Sci USA. 2016;113:E1953–62.

  52. Fouqué A, Lepvrier E, Debure L, Gouriou Y, Malleter M, Delcroix V, et al. The apoptotic members CD95, BclxL, and Bcl-2 cooperate to promote cell migration by inducing Ca(2+) flux from the endoplasmic reticulum to mitochondria. Cell Death Differ. 2016;23:1702–16.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  53. Shimizu S, Narita M, Tsujimoto Y. Bcl-2 family proteins regulate the release of apoptogenic cytochrome c by the mitochondrial channel VDAC. Nature. 1999;399:483–7.

    Article  CAS  PubMed  Google Scholar 

  54. Rostovtseva TK, Tan W, Colombini M. On the role of VDAC in apoptosis: fact and fiction. J Bioenerg Biomembr. 2005;37:129–42.

    Article  CAS  PubMed  Google Scholar 

  55. Tan W, Colombini M. VDAC closure increases calcium ion flux. Biochim Biophys Acta. 2007;1768:2510–5.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Popgeorgiev N, Bonneau B, Ferri KF, Prudent J, Thibaut J, Gillet G. The apoptotic regulator Nrz controls cytoskeletal dynamics via the regulation of Ca2+ trafficking in the zebrafish blastula. Dev Cell. 2011;20:663–76.

    Article  CAS  PubMed  Google Scholar 

  57. Mookerjee SA, Gerencser AA, Nicholls DG, Brand MD. Quantifying intracellular rates of glycolytic and oxidative ATP production and consumption using extracellular flux measurements. J Biol Chem. 2017;292:7189–207.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Suzuki J, Kanemaru K, Ishii K, Ohkura M, Okubo Y, Iino M. Imaging intraorganellar Ca2+ at subcellular resolution using CEPIA. Nat Commun. 2014;5:4153.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We would like to thank Stéphane Borel for technical assistance, Julien Prudent for critical reading of the manuscript, Brigitte Manship for manuscript editing, Isabelle Goddard for skilful technical assistance regarding studies in mice (LMT, Lyon), and CIQLE core facility (SFR Santé Lyon-Est) for videomicroscopy. This work was supported by AFM telethon (to GG, grant # 20269) and Fondation ARC to GG (grant # PGA1 RF20180206799) and JL (grant # PJA 20161204606).

Author information

Author notes

  1. Mathieu Deygas

    Present address: Institut Curie, CNRS UMR144, 6 Rue Jean Calvin, 75005, Paris, France

  2. Adrien Nougarède

    Present address: CEA LETI, 17 Avenue des Martyrs, 38000, Grenoble, France

Authors and Affiliations

  1. Université de Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, 69008, Lyon, France

    Margaux Bessou, Jonathan Lopez, Rudy Gadet, Mathieu Deygas, Nikolay Popgeorgiev, Delphine Poncet, Adrien Nougarède, Pauline Billard, Ivan Mikaelian, Philippe Gonzalo, Ruth Rimokh & Germain Gillet

  2. Hospices Civils de Lyon, Centre de Biologie Sud, Centre Hospitalier Lyon Sud, Chemin du Grand Revoyet, 69495, Pierre Bénite, France

    Jonathan Lopez & Delphine Poncet

  3. Laboratoire de Biochimie, CHU de Saint-Etienne, 42000 Saint-Etienne, France

    Philippe Gonzalo

  4. Hospices Civils de Lyon, Laboratoire d’Anatomie et Cytologie Pathologiques, Centre Hospitalier Lyon Sud, Chemin du Grand Revoyet, 69495, Pierre Bénite, France

    Germain Gillet

Authors
  1. Margaux Bessou
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jonathan Lopez
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Rudy Gadet
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Mathieu Deygas
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Nikolay Popgeorgiev
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Delphine Poncet
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Adrien Nougarède
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Pauline Billard
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Ivan Mikaelian
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Philippe Gonzalo
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Ruth Rimokh
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Germain Gillet
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Ruth Rimokh or Germain Gillet.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

These authors contributed equally: Ruth Rimokh, Germain Gillet

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Bessou, M., Lopez, J., Gadet, R. et al. The apoptosis inhibitor Bcl-xL controls breast cancer cell migration through mitochondria-dependent reactive oxygen species production. Oncogene 39, 3056–3074 (2020). https://doi.org/10.1038/s41388-020-1212-9

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41388-020-1212-9

This article is cited by

Search

Advanced search

Quick links