Divergent Evolution of Human p53 Binding Sites: Cell Cycle Versus Apoptosis

Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, United States of America.
PLoS Genetics (Impact Factor: 7.53). 07/2007; 3(7):e127. DOI: 10.1371/journal.pgen.0030127
Source: PubMed


The p53 tumor suppressor is a sequence-specific pleiotropic transcription factor that coordinates cellular responses to DNA damage and stress, initiating cell-cycle arrest or triggering apoptosis. Although the human p53 binding site sequence (or response element [RE]) is well characterized, some genes have consensus-poor REs that are nevertheless both necessary and sufficient for transactivation by p53. Identification of new functional gene regulatory elements under these conditions is problematic, and evolutionary conservation is often employed. We evaluated the comparative genomics approach for assessing evolutionary conservation of putative binding sites by examining conservation of 83 experimentally validated human p53 REs against mouse, rat, rabbit, and dog genomes and detected pronounced conservation differences among p53 REs and p53-regulated pathways. Bona fide NRF2 (nuclear factor [erythroid-derived 2]-like 2 nuclear factor) and NFkappaB (nuclear factor of kappa light chain gene enhancer in B cells) binding sites, which direct oxidative stress and innate immunity responses, were used as controls, and both exhibited high interspecific conservation. Surprisingly, the average p53 RE was not significantly more conserved than background genomic sequence, and p53 REs in apoptosis genes as a group showed very little conservation. The common bioinformatics practice of filtering RE predictions by 80% rodent sequence identity would not only give a false positive rate of approximately 19%, but miss up to 57% of true p53 REs. Examination of interspecific DNA base substitutions as a function of position in the p53 consensus sequence reveals an unexpected excess of diversity in apoptosis-regulating REs versus cell-cycle controlling REs (rodent comparisons: p < 1.0 e-12). While some p53 REs show relatively high levels of conservation, REs in many genes such as BAX, FAS, PCNA, CASP6, SIVA1, and P53AIP1 show little if any homology to rodent sequences. This difference suggests that among mammalian species, evolutionary conservation differs among p53 REs, with some having ancient ancestry and others of more recent origin. Overall our results reveal divergent evolutionary pressure among the binding targets of p53 and emphasize that comparative genomics methods must be used judiciously and tailored to the evolutionary history of the targeted functional regulatory regions.

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Available from: Monica M Horvath
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    • "Heiber and Barber (2011) who demonstrated that VSV encoding mouse p53 is not attenuated in vitro, but highly attenuated in normal tissues in vivo, and that in vivo treatment of mice results in enhanced tumor killing through stimulated antitumor immunity (Heiber and Barber, 2011). It should be noted that analysis of murine and human p53 response element consensus sequences show many differences between murine and human p53, suggesting that caution should be taken when generalizing about the similarity of regulation of the p53 pathway between humans and rodents (Horvath et al., 2007). Nevertheless, our present data and the previous study using VSV encoding mouse p53 in vivo (Heiber and Barber, 2011) both show VSV-p53 as a promising OV. "
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    ABSTRACT: Virus-encoded tumor suppressor p53 transgene expression has been successfully used in vesicular stomatitis virus (VSV) and other oncolytic viruses (OVs) to enhance their anticancer activities. However, p53 is also known to inhibit virus replication via enhanced type I interferon (IFN) antiviral responses. To examine whether p53 transgenes enhance antiviral signaling in human pancreatic ductal adenocarcinoma (PDAC) cells, we engineered novel VSV recombinants encoding human p53 or the previously described chimeric p53-CC, which contains the coiled-coil (CC) domain from breakpoint cluster region (BCR) protein and evades the dominant-negative activities of endogenously expressed mutant p53. Contrary to an expected enhancement of antiviral signaling by p53, our global analysis of gene expression in PDAC cells showed that both p53 and p53-CC dramatically inhibited type I IFN responses. Our data suggest that this occurs through p53-mediated inhibition of the NF-κB pathway. Importantly, VSV-encoded p53 or p53-CC did not inhibit antiviral signaling in non-malignant human pancreatic ductal cells, which retained their resistance to all tested VSV recombinants. To the best of our knowledge, this is the first report of p53-mediated inhibition of antiviral signaling, and it suggests that OV-encoded p53 can simultaneously produce anticancer activities while assisting, rather than inhibiting, virus replication in cancer cells. Copyright © 2015 Elsevier Inc. All rights reserved.
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    • "Although thousands of p53 binding sites have been mapped in vivo based on chromatin immunoprecipitation (11–13), only ∼150 response elements (REs) are proven to be functional (4). From the evolutionary point of view, it is remarkable that the human CCA-sites display high similarity with their counterparts in the mouse and rat genomes, while the Apo-sites as a group show little homology to the corresponding rodent sequences (14,15). Consistent with this observation, we found that 13 out of ∼150 human p53 REs (4) occur in primate-specific Alu repeats and 10 of them belong to the Apo-sites (16). "
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    ABSTRACT: The tumor suppressor protein p53 exhibits high affinity to the response elements regulating cell cycle arrest genes (CCA-sites), but relatively low affinity to the sites associated with apoptosis (Apo-sites). This in vivo tendency cannot be explained solely by the p53-DNA binding constants measured in vitro. Since p53 can bind nucleosomal DNA, we sought to understand if the two groups of p53 sites differ in their accessibility when embedded in nucleosomes. To this aim, we analyzed the sequence-dependent bending anisotropy of human genomic DNA containing p53 sites. For the 20 CCA-sites, we calculated rotational positioning patterns predicting that most of the sites are exposed on the nucleosomal surface. This is consistent with experimentally observed positioning of human nucleosomes. Remarkably, the sequence-dependent DNA anisotropy of both the p53 sites and flanking DNA work in concert producing strong positioning signals. By contrast, both the predicted and observed rotational settings of the 38 Apo-sites in nucleosomes suggest that many of these sites are buried inside, thus preventing immediate p53 recognition and delaying gene induction. The distinct chromatin organization of the CCA response elements appears to be one of the key factors facilitating p53-DNA binding and subsequent activation of genes associated with cell cycle arrest.
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    • "The overlap between the two main data sources is analyzed in Supplementary Table S1. To avoid overfitting when building the model, we included eight validated p53 binding sites from (29) that differed from the known p53 consensus sequence (Supplementary Table S2). For the ChIP-PET data (28), we extracted the hg17 coordinates of all clusters with three or more overlapping DNA fragments (PET-3+ clusters) from UCSC ( "
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    ABSTRACT: Genome-wide prediction of transcription factor binding sites is notoriously difficult. We have developed and applied a logistic regression approach for prediction of binding sites for the p53 transcription factor that incorporates sequence information and chromatin modification data. We tested this by comparison of predicted sites with known binding sites defined by chromatin immunoprecipitation (ChIP), by the location of predictions relative to genes, by the function of nearby genes and by analysis of gene expression data after p53 activation. We compared the predictions made by our novel model with predictions based only on matches to a sequence position weight matrix (PWM). In whole genome assays, the fraction of known sites identified by the two models was similar, suggesting that there was little to be gained from including chromatin modification data. In contrast, there were highly significant and biologically relevant differences between the two models in the location of the predicted binding sites relative to genes, in the function of nearby genes and in the responsiveness of nearby genes to p53 activation. We propose that these contradictory results can be explained by PWM and ChIP data reflecting primarily biophysical properties of protein–DNA interactions, whereas chromatin modification data capture biologically important functional information.
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