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Ceramide induces mitochondrial activation and apoptosis via a Bax-dependent pathway in human carcinoma cells

Oncogene volume 21, pages 4009–4019 (2002)Cite this article

Abstract

The intracellular pathways leading to mitochondrial activation and subsequent cell death in the ceramide-mediated stress response have been intensively studied in recent years. Experimental evidence has been provided that ceramide-induced apoptosis is inhibited by overexpression of antiapoptotic proteins of the Bcl-2 family. However, the direct effect of proapoptotic gene products, e.g. Bax, on ceramide-induced death signalling has not yet been studied in detail. In the present work, we show by measurement of mitochondrial permeability transition, cytochrome c release, activation of caspase-3 and DNA fragmentation that ceramide-induced apoptosis is marginal in Bax-negative DU 145 cells. Reconstitution of Bax by generation of DU 145 cells stably expressing this proapoptotic factor, clearly enhanced ceramide-induced apoptosis at all levels of the mitochondrial signalling cascade. Using the broad-range caspase inhibitor zVAD-fmk and zDEVD-fmk, an inhibitor of caspase-3-like activities, we demonstrate that the ceramide-induced mitochondrial activation in Bax-transfected DU 145 cells is caspase-independent. On the other hand, apoptotic events located downstream of the mitochondria, e.g. DNA fragmentation, were shown to be caspase-dependent. This influence of Bax on ceramide-induced apoptosis was confirmed in another cellular system: whereas Bax-positive HCT116 wild type cells were very sensitive towards induction of cell death by C2-ceramide, sensitivity of Bax knock-out HCT116 cells was significantly reduced. Thus, we conclude that Bax is a key activator of ceramide-mediated death pathways.

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Acknowledgements

This work was supported by grants from the Deutsche Forschungsgemeinschaft (SFB 506) to PT Daniel, the European Community TMR Programme to PT Daniel, the Deutsche José Carreras Leukämie-Stiftung e. V. (DJCLS-R01/02) to PT Daniel and T Wieder, the Verein zur Förderung der Tagesklinik e. V. and the Berliner Krebsgesellschaft e. V. to T Wieder. The authors want to thank Bert Vogelstein (Johns Hopkins University School of Medicine, Baltimore, MD, USA) for kindly providing HCT116 wild type and Bax −/− cells. The excellent technical assistance of Verena Lehmann is gratefully acknowledged.

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  1. Clarissa von Haefen and Thomas Wieder: C von Haefen and T Wieder contributed equally to this study

Authors and Affiliations

  1. Department of Hematology, Oncology and Tumor Immunology, University Medical Center Charité, Humboldt University of Berlin, Berlin, 13125, Germany

    Clarissa von Haefen, Thomas Wieder, Bernd Gillissen, Lilian Stärck, Vilma Graupner, Bernd Dörken & Peter T Daniel

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  1. Clarissa von Haefen
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Correspondence to Peter T Daniel.

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von Haefen, C., Wieder, T., Gillissen, B. et al. Ceramide induces mitochondrial activation and apoptosis via a Bax-dependent pathway in human carcinoma cells. Oncogene 21, 4009–4019 (2002). https://doi.org/10.1038/sj.onc.1205497

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