Abstract
The role of ceramide in triggering apoptosis is still a matter of debate. While in some experimental systems, ceramide was shown to mediate Fas-induced cell death, in other instances it was claimed to induce the expression of Fas ligand (FasL), killing cells in a caspase-dependent fashion. We found that, in mature A20 B cells, ceramide-induced apoptosis is independent of the caspase pathway, since we observed no ICE-like, CPP32-like and Mch2 activities and no PARP proteolysis. Moreover, we were unable to protect these cells from ceramide-induced apoptosis using caspase inhibitors, while they blocked Fas-induced apoptosis and no FasL induction could be detected following ceramide treatment. These results suggest that ceramide does not induce apoptosis through the Fas/FasL pathway. We also found that overexpression of Nur77, a zinc-finger transcription factor described to upregulate FasL, antagonizes ceramide-induced apoptosis, but not Fas-induced apoptosis. This further supports the hypothesis that Fas and ceramide death pathways are independent in A20 cells. Ceramide-induced cell death was associated with increased c-myc, p53, Bax and p27kip1 levels; in contrast, cells transfected with Nur77 (A20Nur77), resistant to ceramide-induced apoptosis, showed a marked downregulation of p53 after ceramide treatment, with neither Bax nor p27kip1 induction. In conclusion, our results suggest that, in the A20 B cell line, Fas and ceramide trigger two distinct pathways and that Nur77 overexpression confers protection against ceramide-mediated apoptosis which correlates with inhibition of p53, Bax and p27kip1 induction.
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Abbreviations
- ICE:
-
interleukin-1β converting enzyme
- PARP:
-
poly(ADP-ribose) polymerase
- PCD:
-
programmed cell death
- CrmA:
-
cytokine response modifier A
- TNF:
-
tumor necrosis factor
- Ac-YVAD-CMK:
-
acetyl-Tyr-Val-Ala-Asp-chloromethylketone
- Ac-DEVD-CHO:
-
acetyl-Asp-Glu-Val-Asp-aldehyde
- Z-VAD-fmk:
-
Z-Val-Ala-DL-Asp-fluoromethylketone
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Acknowledgements
We thank Drs. Lester F Lau for pSG5/Nur77 and M Serrano for p21waf1/cip1 and p27kip1 antibodies and Drs. R Geha and M Serrano for critical reading of the manuscript. We also thank Drs. I Palmero and I Flores for helpful comments and discussion and C Mark for editorial assistance. This work was supported by grants from the Dirección General de Ciencia y Tecnología (DGCyT) and the European Union. The Department of Immunology and Oncology was founded and is supported by the CSIC and Pharmacia & Upjohn.
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Brás, A., Albar, J., Leonardo, E. et al. Ceramide-induced cell death is independent of the Fas/Fas ligand pathway and is prevented by Nur77 overexpression in A20 B cells. Cell Death Differ 7, 262–271 (2000). https://doi.org/10.1038/sj.cdd.4400653
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DOI: https://doi.org/10.1038/sj.cdd.4400653
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