p53-dependent down-regulation of telomerase is mediated by p21waf1.
Inactivation of p53 and activation of telomerase occur in the majority of human cancers, raising the possibility of a link between these two pathways. Overexpression of wild-type p53 down-regulates the enzymatic activity of telomerase in various cancer cell lines through transcriptional repression of its catalytic subunit, human telomerase reverse transcriptase (hTERT). In this study, we re-evaluated the role of p53 in telomerase regulation using isogenic cell lines expressing physiological levels of p53. We demonstrate that endogenous wild-type p53 was able to down-regulate telomerase activity, hTERT mRNA levels, and promoter activity; however, the ability to repress hTERT expression was found to be cell type-specific. The integrity of the DNA-binding core domain, the N-terminal transactivation domain, and the C-terminal oligomerization domains of p53 was essential for hTERT promoter repression, whereas the proline-rich domain and the extreme C terminus were not required. Southwestern and chromatin immunoprecipitation experiments demonstrated lack of p53 binding to the hTERT promoter, raising the possibility of an indirect repressive mechanism. The down-regulation of hTERT promoter activity was abolished by a dominant-negative E2F1 mutant. Mutational analysis identified a specific E2F site responsible for p53-mediated repression. Knockdown of the key p53 transcriptional target, p21, was sufficient to eliminate the p53-dependent repression of hTERT. Inactivation of the Rb family using either viral oncoproteins or RNA interference attenuated the repression. Inhibition of histone deacetylases also interfered with the repression of hTERT by p53. Therefore, our results suggest that repression of hTERT by endogenous p53 is mediated by p21 and E2F.
Shats
Igor
I
Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot 76100, Israel.
Milyavsky
Michael
M
Tang
Xiaohu
X
Stambolsky
Perry
P
Erez
Neta
N
Brosh
Ran
R
Kogan
Ira
I
Braunstein
Ilana
I
Tzukerman
Maty
M
Ginsberg
Doron
D
Rotter
Varda
V
D001483:Base Sequence; D015139:Blotting, Southern; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004247:DNA / Q000737:chemistry; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism; D015536:Down-Regulation; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D017930:Genes, Reporter; D006655:Histone Deacetylases / Q000378:metabolism; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D009369:Neoplasms / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011392:Proline / Q000737:chemistry; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019098:Telomerase / Q000096:biosynthesis*; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism / Q000502:physiology*
C493956:CDKN1A protein, human; D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D012333:RNA, Messenger; D016160:Retinoblastoma Protein; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D004247:DNA; D011392:Proline; D008156:Luciferases; D019098:Telomerase; D006655:Histone Deacetylases