Abstract
The mechanism of p53-dependent apoptosis is still only partly defined. Using early-passage embryonic fibroblasts (MEF) from wild-type (wt), p53−/− and bax−/− mice, we observe a p53-dependent translocation of Bax to the mitochondria and a release of mitochondrial Cytochrome c during stress-induced apoptosis. These events proceed independent of zVAD-inhibitable caspase activation, are not prevented by dominant negative FADD (DN-FADD), but are negatively regulated by Mdm-2. Bcl-xL expression prevents the release of mitochondrial Cytochrome c and apoptosis, but not Bax translocation. At a single-cell level, enforced expression of p53 is sufficient to induce Bax translocation and Cytochrome c release. Real-time RT-PCR analysis reveals a significant induction of RNA expression of Noxa and Bax in p53+/+, but not in p53−/− MEF. Noxa protein expression becomes detectable prior to Bax translocation, and downregulation of endogenous Noxa by RNA interference protects wt MEF against p53-dependent apoptosis. Hence, in oncogene-expressing MEF p53 induces apoptosis by BH3 protein-dependent caspase activation.
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Abbreviations
- DMEM:
-
Dulbecco's modified essential medium
- FADD:
-
Fas-associated death domain
- MEF:
-
murine embryonic fibroblasts
- PARP:
-
poly(ADP-ribose) polymerase
- PCR:
-
polymerase chain reaction
- RNAi:
-
RNA interference
- TMRE:
-
tetramethyl-rhodamine-ethylester
- zVAD-fmk:
-
benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone
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Acknowledgements
We are indebted to Dr. SJ Korsmeyer for providing cells from Bax-deficient mice, to Dr. T Lin for animal care and to P Fitzgerald for help with cloning. Drs. SW Lowe, GJ Hannon, T Kitamura, M Oren, S Hedrick, DC Maneval, RJ Youle and R Agami are thanked for providing expression vectors.
This work was supported by National Institutes of Health grants GM52735, CA69381, and AI40646 awarded to DRG, and by the Max-Eder-Programm of the Deutsche Krebshilfe to MS (70-2952).
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Schuler, M., Maurer, U., Goldstein, J. et al. p53 triggers apoptosis in oncogene-expressing fibroblasts by the induction of Noxa and mitochondrial Bax translocation. Cell Death Differ 10, 451–460 (2003). https://doi.org/10.1038/sj.cdd.4401180
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DOI: https://doi.org/10.1038/sj.cdd.4401180
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