Mutant p53 (mtp53) promotes chemotherapy resistance through multiple mechanisms, including disabling proapoptotic proteins and regulating gene expression. Comparison of genome wide analysis of mtp53 binding revealed that the ETS-binding site motif (EBS) is prevalent within predicted mtp53-binding sites. We demonstrate that mtp53 regulates gene expression through EBS in promoters and that ETS2 mediates the interaction with this motif. Importantly, we identified TDP2, a 59-tyrosyl DNA phosphodiesterase involved in the repair of DNA damage caused by etoposide, as a transcriptional target of mtp53. We demonstrate that suppression of TDP2 sensitizes mtp53-expressing cells to etoposide and that mtp53 and TDP2 are frequently overexpressed in human lung cancer; thus, our analysis identifies a potentially ''druggable'' component of mtp53's gain-of-function activity.[Keywords: TDP2; cancer; p53] Supplemental material is available for this article. One of the definitive characteristics of the mutant p53 (mtp53) protein is that it can alter the cellular phenotype, resulting in the acquisition of gain-of-function activities such as abnormal cell growth, suppression of apoptosis, chemotherapy resistance, increased angiogenesis, and metastasis ( For example, mtp53 can interact with its family members, p63 and p73, and disable their ability to induce apoptosis (Di Como et al. 1999;Marin et al. 2000;Strano et al. 2000Strano et al. , 2002Gaiddon et al. 2001;Bergamaschi et al. 2003;Irwin et al. 2003;Lang et al. 2004). mtp53 can also interact with other transcription factors (such as NF-Y, E2F1, VDR, and p63) and thereby can be recruited to target genes that have consensus binding sites for these transcription factors (Di Agostino et al. 2006;Adorno et al. 2009;Fontemaggi et al. 2009;Stambolsky et al. 2010). Notably, some of these interactions help explain how the mtp53 protein can deregulate gene expression and promote abnormal cell growth, angiogenesis, and metastasis (Di Agostino et al. 2006;Adorno et al. 2009;Fontemaggi et al. 2009;Muller et al. 2009Muller et al. , 2011. However, thus far, none of these transcription factors have been shown to play a fundamental role in regulating the expression of genes that can confer chemotherapy resistance by modulating the response to DNA damage. The main goal of this study was to identify a transcriptional regulatory mechanism through which mtp53 can promote chemotherapy resistance.
Results
Identification of mtp53 target genesTo identify transcriptional targets of mtp53, we employed two different approaches: chromatin immunoprecipitation (ChIP)-on-chip and ChIP combined with deep sequencing (ChIP-seq). The ChIP-on-chip was performed with Nimblegen arrays that have oligonucleotide probes for all of the promoters in the human genome (Nimblegen Promoter Arrays). The ChIP-seq analysis was performed using the Illumina platform. We conducted these analyses in the Li-Fraumeni cell line MDAH087, which expresses only the R248W mtp53 protein (Bischoff et al. 1990). The ChIP-on-chip analysis identif...
