Whole-genome cartography of p53 response elements ranked on transactivation potential

Background Many recent studies using ChIP-seq approaches cross-referenced to trascriptome data and also to potentially unbiased in vitro DNA binding selection experiments are detailing with increasing precision the p53-directed gene regulatory network that, nevertheless, is still expanding. However, most experiments have been conducted in established cell lines subjected to specific p53-inducing stimuli, both factors potentially biasing the results. Results We developed p53retriever, a pattern search algorithm that maps p53 response elements (REs) and ranks them according to predicted transactivation potentials in five classes. Besides canonical, full site REs, we developed specific pattern searches for non-canonical half sites and 3/4 sites and show that they can mediate p53-dependent responsiveness of associated coding sequences. Using ENCODE data, we also mapped p53 REs in about 44,000 distant enhancers and identified a 16-fold enrichment for high activity REs within those sites in the comparison with genomic regions near transcriptional start sites (TSS). Predictions from our pattern search were cross-referenced to ChIP-seq, ChIP-exo, expression, and various literature data sources. Based on the mapping of predicted functional REs near TSS, we examined expression changes of thirteen genes as a function of different p53-inducing conditions, providing further evidence for PDE2A, GAS6, E2F7, APOBEC3H, KCTD1, TRIM32, DICER, HRAS, KITLG and TGFA p53-dependent regulation, while MAP2K3, DNAJA1 and potentially YAP1 were identified as new direct p53 target genes. Conclusions We provide a comprehensive annotation of canonical and non-canonical p53 REs in the human genome, ranked on predicted transactivation potential. We also establish or corroborate direct p53 transcriptional control of thirteen genes. The entire list of identified and functionally classified p53 REs near all UCSC-annotated genes and within ENCODE mapped enhancer elements is provided. Our approach is distinct from, and complementary to, existing methods designed to identify p53 response elements. p53retriever is available as an R package at: http://tomateba.github.io/p53retriever. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-1643-9) contains supplementary material, which is available to authorized users.


-Supplementary Text
Information on nine recently proposed or established direct p53 target genes with mapped functional p53 REs that were confirmed as p53-responsive by qPCR (see Figure 5A).

PDE2A
PDE2A is a cyclic nucleotide (cGMP and cAMP) phosphodiesterase that has been reported as potential marker of cancer treatments [1] and up-regulated in -catenin mutated adrenocortical carcinomas [2]. Inhibition of PDE2A activity in melanoma cells correlated with G2/M arrest [3]. A PDE2A splice variant was found capable to localize in mitochondria and to regulate respiration [4].

GAS6
GAS6 is a ligand for tyrosine kinase TAM receptors, including AXL. Previously, GAS6 and other genes in a linkage cluster were found overexpressed in aggressive mammary tumors from a p53-null mouse model [5]. Further, high GAS6 expression was an adverse prognostic marker in adult AML patients [6]. High expression of CXCL12 and CXCR4, potentially involved in tumor metastasis [7] was associated with GAS6 expression.
However, activated TAM receptors can have both pro-and anti-inflammatory effects, depending on the cell type [8]. As for PDE2A, the physiological role of p53-dependent control of GAS6 expression need to be elucidated.

E2F7
E2F7 is an inhibitory member of the E2F family of transcription factor that does not contain a RB1 binding pocket [9] and can compensate for the loss of RB1 acting by repressing cell cycle progression. p53-dependent activation of E2F7 and a role in controlling proliferation and inducing senescence has been recently reported [10,11].

APOBEC3H
APOBEC3H belongs to the APOBEC3 family of DNA deaminases, with important role in antiviral defense and also in retro-transposon mobility. APOBEC3C but not 3H was recently discovered as an important mediator of hypermutability in somatic cancer via DNA editing [12]. A role for p53 in the inhibition of retroposition and in counteracting viral infection has been establish and the control of APOBEC3H expression could play an important role in this processes.

KCTD1
KCTD1 is a nuclear protein that functions as a transcriptional repressor. Very recently it was shown to enhance -catenin degradation, thus suppressing Wnt-mediated signaling [13]. As germline mutations in this gene have been associated with a form of ectodermal dyplasias [14], it would be interesting to assess if KCTD1 can also be a direct target of the p63 transcription factor.

TRIM32
TRIM32 is an E3-ubiquitin ligase protein member of a family that plays a role in HIV infectivity [15], but it has been linked to distinct phenotypes in cancer cells including promotion of apoptosis [16] and differentiation [17].

TGFA
This gene encodes a growth factor that is a ligand for the epidermal growth factor receptor (EGFR), which activates a signaling pathway for cell proliferation, differentiation and development. Moreover, it has been involved in cancer progression. Recent data identified TGFA and other growth factors as major players in the control of anti-apoptotic activities in p53 negative cells. For instance, PDGFRB, IGFR1R and TGFA are upregulated in HCT116 p53 −/− cells compared to the HCT116 p53 +/+ [18].

HRAS
This gene belongs to the Ras oncogene family whose members can bind GTP and GDP, and have intrinsic GTPase activity. These family members are crucial players in many signaling networks that regulate cell-cycle progression, growth, migration, cytoskeletal changes, apoptosis, and senescence [19].