The binding properties and biological activities of Bcl-2 and Bax in cells exposed to apoptotic stimuli.
The oncogene product Bcl-2 protects cells from apoptosis whereas its homolog Bax functions to kill cells. Several binding partners of Bcl-2 and Bax have been isolated, but none of them has yet provided clues as to exactly how Bcl-2 and Bax work. According to one view, Bcl-2 and Bax interact with survival and death effector molecules, respectively, and neutralize each other through heterodimerization. Alternatively, Bcl-2 requires Bax for death protection, and additional proteins bind to the heterodimer to regulate its activity. Here we used a co-immunoprecipitation strategy to distinguish between these two possibilities. We show that the Bcl-2-Bax heterodimer is maintained, and no other protein associates stably in detectable amounts with Bcl-2, Bax, or the heterodimer in anti-Bcl-2 and anti-Bax immunoprecipitates from normal cells and cells exposed to apoptotic stimuli. Analysis of cells expressing various levels of Bcl-2 and Bax, however, revealed that the degree of protection against apoptosis does not correlate with the number of Bcl-2-Bax heterodimers but the amount of Bcl-2 that is free of Bax. In addition, the survival activity of Bcl-2 is unaffected when Bax expression is ablated by an antisense strategy. Our findings suggest that the Bcl-2-Bax heterodimer is a negative regulator of death protection, and that Bcl-2 requires neither Bax nor major, stable interactions with other cellular proteins to exert its survival function. We therefore propose that Bcl-2 acts as an enzyme (capturing substrates in a transient way), as a homodi- or multimer, or through the interaction with non-proteaceous targets (lipids, ions).
Otter
I
I
Institute of Biochemistry, University of Fribourg, Rue du Musée 5, CH-1700 Fribourg, Switzerland.
Conus
S
S
Ravn
U
U
Rager
M
M
Olivier
R
R
Monney
L
L
Fabbro
D
D
Borner
C
C
D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D019281:Dimerization; D007976:Leupeptins / Q000494:pharmacology; D051379:Mice; D011233:Precipitin Tests; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D019311:Staurosporine / Q000494:pharmacology; D051028:bcl-2-Associated X Protein
C494827:Bax protein, mouse; C494828:Bax protein, rat; D007976:Leupeptins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011994:Recombinant Proteins; D051028:bcl-2-Associated X Protein; D019311:Staurosporine; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde