Inhibition of p53 transcriptional activity by Bcl-2 requires its membrane-anchoring domain.
We show here that the anti-apoptosis protein Bcl-2 potently inhibits p53-dependent transcriptional activation of various p53-responsive promoters in reporter gene co-transfection assays in human embryonic kidney 293 and MCF7 cells, without affecting nuclear accumulation of p53 protein. In contrast, Bcl-2(Deltatransmembrane (TM)), which lacks a hydrophobic membrane-anchoring domain, had no effect on p53 activity. Similarly, in MCF7 cells stably expressing either Bcl-2 or Bcl-2(DeltaTM), nuclear levels of p53 protein were up-regulated upon treatment with the DNA-damaging agents doxorubicin and UV radiation, whereas p53-responsive promoter activity and expression of p21(CIP1/WAF1) were strongly reduced in MCF7-Bcl-2 cells but not in MCF7-Bcl-2(DeltaTM) or control MCF7 cells. The issue of membrane anchoring was further explored by testing the effects of Bcl-2 chimeric proteins that contained heterologous transmembrane domains from the mitochondrial protein ActA or the endoplasmic reticulum protein cytochrome b5. Both Bcl-2(ActA) and Bcl-2(Cytob5) suppressed p53-mediated transactivation of reporter gene plasmids with efficiencies comparable to wild-type Bcl-2. These results suggest that (a) Bcl-2 not only suppresses p53-mediated apoptosis but also interferes with the transcriptional activation of p53 target genes at least in some cell lines, and (b) membrane anchoring is required for this function of Bcl-2. We speculate that membrane-anchored Bcl-2 may sequester an unknown factor necessary for p53 transcriptional activity.
Froesch
B A
BA
Burnham Institute, La Jolla, California 92037, USA.
Aimé-Sempé
C
C
Leber
B
B
Andrews
D
D
Reed
J C
JC
D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics* / Q000528:radiation effects; D002460:Cell Line; D004249:DNA Damage / Q000187:drug effects / Q000528:radiation effects; D004317:Doxorubicin / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000528:radiation effects; D006801:Humans; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D015533:Transcriptional Activation; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis / Q000235:genetics*; D014466:Ultraviolet Rays
D000970:Antineoplastic Agents; D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin