Abstract A53: Dose-dependent levels of p53 mediate different cell fate decisions in human mammary epithelial cells

Abstract

Abstract Introduction: The p53 protein is a tumor suppressor which is activated in response to diverse cellular stress signals such as DNA damage or abnormal oncogene activation. Active p53 triggers various anti-proliferative programs primarily through its ability to transcriptionally activate target genes that mediate these processes. p53 can induce cell cycle arrest, senescence or apoptosis, but the exact mechanisms involved in determining these cell fate decisions remain elusive. A p53 dose dependent model has been put forward to explain the choice of cell fate upon p53 expression. According to this model, low expression levels of p53 induce cell cycle arrest, while high expression levels of p53 trigger cell death. p53-dependent cell cycle arrest or apoptosis could be mediated by the same target genes induced to lower or higher levels, respectively. Alternatively, different levels of p53 could induce distinct sets of target genes with different biological outcomes. Methods: To investigate the role of p53 in cell fate decisions, we developed a doxycycline-regulatable lentiviral system which allows tight regulation of p53 expression in a titratable, dose-dependent manner. Using this system, we have generated a mass population of non-tumorigenic B5/589 human mammary epithelial cells that inducibly expresses p53 upon doxycycline addition. Results: We identified physiological levels of p53 that were capable of inducing p53-dependent cell cycle arrest or cell death within the same mass population. Our data demonstrated that a narrow window of low p53 expression induced G1 cell cycle arrest without detectable apoptosis. Above that level, cells underwent apoptosis. These results suggested that a threshold level of p53 expression can be a major deciding factor between these two cell fates. To compare the transcriptional programs leading to p53-dependent arrest or apoptosis, we conducted a whole-transcript microarray analysis using Human GeneChip 1.0 ST Array System (Affymetrix) in p53-inducible B5/589 cells. Using linear models for microarray data (LIMMA), our preliminary analysis identified 251 and 610 genes upregulated after 12h of low and high p53 induction; and 277 and 860 genes upregulated after 24h of low and high p53 induction, respectively. 259 genes were significantly upregulated following 24h of high versus low p53 induction. Functional Annotation Enrichment analysis of the 259 genes detected a highly significant subset of genes involved in the induction of apoptosis. Of note, 117 genes were specifically downregulated following 24h of high versus low p53 induction. Further analysis will be focused on the identification of pathways specifically engaged during cell cycle arrest and apoptosis. Conclusions: Our results indicate that the level of p53 expression plays an important role in cell fate decisions between arrest and apoptosis. Furthermore, our microarray data suggest that high physiological levels of p53 regulate a specific subset of target genes associated with p53-dependent cell death. Citation Information: Cancer Res 2009;69(23 Suppl):A53.