p53-dependent repression of the human MCL-1 gene encoding an anti-apoptotic member of the BCL-2 family: the role of Sp1 and of basic transcription factor binding sites in the MCL-1 promoter.
p53 regulates transcription of one anti-apoptotic and four pro-apoptotic members of the BCL-2 family, but nothing is known about the regulation of MCL-1, another antiapoptotic member of this family, by p53. Confocal microscopic analysis of COS1, HEK 293 and HeLa cells transfected with a p53 expression plasmid demonstrated a decrease in the signal of endogenous MCL-1 compared to neighboring non-transfected cells. Transcription regulation assays showed that the 1826 bp human MCL-1 promoter fragment was repressed up to 30-fold by wild-type p53 in a dose-dependent manner. As shown by electrophoretic mobility shift assays, Sp1 binding to the sites located in the -295 to +16 MCL-1 promoter fragment was decreased in the presence of p53. However, the MCL-1 promoter devoid of all Sp1 binding sites was still repressed by p53, albeit 2-fold weaker than the wild-type promoter. Overexpression of Sp1 reduced p53-dependent repression of the MCL-1 promoter only up to 2.2-fold. Transcription regulation assays performed with MCL-1 promoter deletion mutants showed that most of the p53 inhibitory effect was mediated by the -41 to +16 bp promoter fragment containing binding sites only for TATA-binding protein and other basal transcription factors. We propose a novel, promoter-based mechanism by which p53 down-regulates expression of the antiapoptotic MCL-1 protein.
Pietrzak
Maciej
M
Department of Endocrinology, Medical Research Center, Polish Academy of Sciences, 02-106 Warsaw, Poland.
Puzianowska-Kuznicka
Monika
M
D000818:Animals; D001665:Binding Sites / Q000235:genetics; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D024202:Electrophoretic Mobility Shift Assay; D006367:HeLa Cells; D006801:Humans; D018613:Microscopy, Confocal; D009154:Mutation; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D035181:TATA-Box Binding Protein / Q000235:genetics / Q000502:physiology; D014158:Transcription, Genetic; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000502:physiology*
D064549:Myeloid Cell Leukemia Sequence 1 Protein; D019253:Proto-Oncogene Proteins c-bcl-2; D016329:Sp1 Transcription Factor; D035181:TATA-Box Binding Protein; D016159:Tumor Suppressor Protein p53