Deconstructing p53 transcriptional networks in tumor suppression

Abstract

p53 is a pivotal tumor suppressor that induces apoptosis, cell-cycle arrest and senescence in response to stress signals. Although p53 transcriptional activation is important for these responses, the mechanisms underlying tumor suppression have been elusive. To date, no single or compound mouse knockout of specific p53 target genes has recapitulated the dramatic tumor predisposition that characterizes p53-null mice. Recently, however, analysis of knock-in mice expressing p53 transactivation domain mutants has revealed a group of primarily novel direct p53 target genes that may mediate tumor suppression in vivo. We present here an overview of well-known p53 target genes and the tumor phenotypes of the cognate knockout mice, and address the recent identification of new p53 transcriptional targets and how they enhance our understanding of p53 transcriptional networks central for tumor suppression.

Figure 1. p53 responds to a plethora of stress signals and regulates diverse responses

Myriad cellular stress signals can activate p53, and p53 can respond by regulating many cellular processes that could contribute to tumor suppression.

Figure 2. Transactivation domains mediate interactions between p53 and cofactors

The domain organization of p53 includes two N-terminal transcriptional activation domains (TADs), a proline rich domain, a DNA binding domain, a tetramerization (Tet) domain, and a basic region. The sequence alignment of mouse and human p53 is shown, with the asterisks indicating the location of the mutations in the TAD mutants. The LW residues mutated in p53^25,26 knock-in mouse strains correspond to amino acids 25 and 26 in mouse p53 and 22 and 23 in human p53, and the FF residues mutated in the p53^53,54 knock-in mouse correspond to amino acids 53 and 54 in both mouse and human p53 (marked in red). A variety of protein cofactors that regulate chromatin remodeling and/or transcriptional initiation interact with p53 via one or both TADs, including the transcriptional regulator proteins TAF6, TAF9, TBP, TRAP80, TFIIH, and histone acetyltransferases p300, CBP, and GCN5, a component of the human STAGA (STP3-TAF(II)31-GCN5-L acetylase) complex. Please note that the exact boundaries of the interaction sites are not precise.

Figure 3. Functional analysis of p53 transactivation domain (TAD) mutants identifies p53 target genes involved in tumor suppression

(a) Transactivation domain 1 (TAD1) and robust transactivation of classical p53 target genes are required for responses to acute DNA damage, including apoptosis and cell cycle arrest. A p53 TAD1 mutant, p53^25,26, is unable to efficiently activate expression of canonical p53 target genes, including p21, Noxa, Perp, and Puma. p53^25,26 is also unable to induce apoptosis or cell cycle arrest in response to acute DNA damage. Transactivation by either TAD1 or TAD2 allows p53 responses to oncogene activation. p53^25,26 can activate expression of only a limited number of mostly novel target genes, but can promote tumor suppression in a number of mouse models. The tumor suppressor capability of p53^25,26 can be explained by its ability to robustly activate a limited set of novel direct p53 target genes (Sidt2, Phlda3, Abhd4, etc.). The capacity of p53^25,26 to promote very low level activation of various classical p53 target genes may also contribute to tumor suppression. The “?” denotes additional, still-unknown genes critical for responses to acute DNA damage and oncogene activation. (b) Comparison of gene expression profiles of p53 wild-type and p53-null HrasV12-MEFs results in more than 1000 differentially expressed genes. To enrich for genes with specific roles in tumor suppression, we leveraged gene expression profiling data generated with the p53^25,26 mutant, which activates only a small subset of p53 target genes, yet is a potent tumor suppressor. Using transcriptomics analysis of HrasV12-MEFs, we identified genes induced at least 2 fold and within 1.5 standard deviations by p53^wt, p53^25,26, and p53^53,54, which all have tumor suppressor activity, relative to p53^25,26,53,54 or p53-null samples, which lack tumor suppressor activity. This list of 130 genes was then filtered for those commonly downregulated in human and mouse tumors, according to the EBI’s Gene Atlas database. A group of 14 candidate genes with likely roles in p53-mediated tumor suppression was defined by this analysis.

Figure 3. Functional analysis of p53 transactivation domain (TAD) mutants identifies p53 target genes involved in tumor suppression

(a) Transactivation domain 1 (TAD1) and robust transactivation of classical p53 target genes are required for responses to acute DNA damage, including apoptosis and cell cycle arrest. A p53 TAD1 mutant, p53^25,26, is unable to efficiently activate expression of canonical p53 target genes, including p21, Noxa, Perp, and Puma. p53^25,26 is also unable to induce apoptosis or cell cycle arrest in response to acute DNA damage. Transactivation by either TAD1 or TAD2 allows p53 responses to oncogene activation. p53^25,26 can activate expression of only a limited number of mostly novel target genes, but can promote tumor suppression in a number of mouse models. The tumor suppressor capability of p53^25,26 can be explained by its ability to robustly activate a limited set of novel direct p53 target genes (Sidt2, Phlda3, Abhd4, etc.). The capacity of p53^25,26 to promote very low level activation of various classical p53 target genes may also contribute to tumor suppression. The “?” denotes additional, still-unknown genes critical for responses to acute DNA damage and oncogene activation. (b) Comparison of gene expression profiles of p53 wild-type and p53-null HrasV12-MEFs results in more than 1000 differentially expressed genes. To enrich for genes with specific roles in tumor suppression, we leveraged gene expression profiling data generated with the p53^25,26 mutant, which activates only a small subset of p53 target genes, yet is a potent tumor suppressor. Using transcriptomics analysis of HrasV12-MEFs, we identified genes induced at least 2 fold and within 1.5 standard deviations by p53^wt, p53^25,26, and p53^53,54, which all have tumor suppressor activity, relative to p53^25,26,53,54 or p53-null samples, which lack tumor suppressor activity. This list of 130 genes was then filtered for those commonly downregulated in human and mouse tumors, according to the EBI’s Gene Atlas database. A group of 14 candidate genes with likely roles in p53-mediated tumor suppression was defined by this analysis.