Abstract
The Bcl-2 family of proteins comprises both cell death inhibiting and cell death promoting members, generally believed to be cytoplasmic and predominantly membrane-associated. Like Bcl-2, many Bcl-2-related proteins contain a C-terminal membrane insertion domain and much research is aimed at evaluating the functional role of their localization to the outer membranes of mitochondria, the endoplasmic reticulum, and perinuclear membranes. However, confocal fluorescence microscopy of human breast cancer cells and rat colon cancer cells immunostained with commercial antibodies raised against different epitopes of the anti-apoptotic Bcl-2 and the pro-apoptotic Bax protein revealed that these proteins are not only present in the cellular cytoplasm, but also within interphase nuclei. This was confirmed by Western blot analysis of isolated nuclei. In human cells, certain epitopes of Bcl-2, but not of Bax, were also found to be associated with mitotic chromatin. Anti-estrogen treatment of human breast cancer cells or transfection with antisense bcl-2 led to a reduction in both cytoplasmic and nuclear Bcl-2. Transfection of human bcl-2 and bax into rat cells resulted in cytoplasmic and nuclear Bcl-2 and Bax. This data seems in line with increasing evidence that the role of the Bcl-2 family of proteins should be extended to activities inside the nuclear compartment.
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Abbreviations
- BC:
-
breast cancer
- BSA:
-
bovine serum albumine
- DAB:
-
diaminobenzidine
- ER:
-
endoplasmic reticulum
- FCS:
-
fetal calf serum
- FITC:
-
fluorescein isothiocyanate
- MAb:
-
monoclonal antibody
- PAb:
-
polyclonal antibodies
- PAGE:
-
polyacrylamide gel electrophoresis
- PARP:
-
poly(ADPribose) polymerase
- PBS:
-
phosphate buffered saline
- PI:
-
propidium iodide
- SDS:
-
sodium dodecyl sulphate
- TM:
-
transmembrane
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Acknowledgements
We would like to thank Hans de Bont and Frans Prins for assisting in confocal laserscan microscopy. The Division of Medical Biochemistry, Sylvius Laboratories, Leiden, is acknowledged for the use of sonification instruments and for advice on subcloning procedures. We thank Joke van Dierendonck-Beukers for critically reading the manuscript and Eduard Bakker for providing the anti-lamin cocktail of antibodies.
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Hoetelmans, R., van Slooten, HJ., Keijzer, R. et al. Bcl-2 and Bax proteins are present in interphase nuclei of mammalian cells. Cell Death Differ 7, 384–392 (2000). https://doi.org/10.1038/sj.cdd.4400664
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DOI: https://doi.org/10.1038/sj.cdd.4400664
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