Refinement of the androgen response element based on ChIP-Seq in androgen-insensitive and androgen-responsive prostate cancer cell lines

Abstract

Sequence motifs are short, recurring patterns in DNA that can mediate sequence-specific binding for proteins such as transcription factors or DNA modifying enzymes. The androgen response element (ARE) is a palindromic, dihexameric motif present in promoters or enhancers of genes targeted by the androgen receptor (AR). Using chromatin immunoprecipitation sequencing (ChIP-Seq) we refined AR-binding and AREs at a genome-scale in androgen-insensitive and androgen-responsive prostate cancer cell lines. Model-based searches identified more than 120,000 ChIP-Seq motifs allowing for expansion and refinement of the ARE. We classified AREs according to their degeneracy and their transcriptional involvement. Additionally, we quantified ARE utilization in response to somatic copy number amplifications, AR splice-variants, and steroid treatment. Although imperfect AREs make up 99.9% of the motifs, the degree of degeneracy correlates negatively with validated transcriptional outcome. Weaker AREs, particularly ARE half sites, benefit from neighboring motifs or cooperating transcription factors in regulating gene expression. Taken together, ARE full sites generate a reliable transcriptional outcome in AR positive cells, despite their low genome-wide abundance. In contrast, the transcriptional influence of ARE half sites can be modulated by cooperating factors.

Genome-wide analysis of AR binding and comparison with transcript expression in primary human fetal prostate fibroblasts and cancer associated fibroblasts

“…Analysis of the binding sequences present in the AR-derived peaks showed that classical palindromic ARE sequences were very poorly represented in fibroblasts, which suggests that the AR is likely to bind to chromatin via cofactors or other transcription factors [20]. AP1 was identified as a co-factor for AR in the PShTERT-AR cell line [34], however we did not observe many AP1 sites in our data.…”

“…This has revealed a transcriptional role for AR in tumour cells regulating cell proliferation, metabolism, survival and DNA repair [18,19]. Prostate cancer cell lines have also been used to determine the changes in AR transcriptional networks in both a hormonenaïve-and castrate-resistant setting and numerous studies have revealed distinct binding profiles in the androgen-independent setting [13,17,18,20,21]. More recently, the genomic binding profile of AR variants have also been explored and have been shown to compensate for full length AR in an endocrine therapy-like setting [22].…”

The Genomic Impact of DNA CpG Methylation on Gene Expression; Relationships in Prostate Cancer

“…In doing so, these consortia have also generated remarkable volumes of publically available data with which to interrogate DNA methylation patterns and relationships to gene expression and cell phenotypes. In cancer it is clear that the TF-genome interactions are corrupted [82,83,84,85] and “re-wired” [86,87,88], for example, by somatic mutations and endogenous structural variants that disrupt TF binding. A major driver of addressing global DNA methylation in cancer has been the development of large cancer genome studies, for example the Cancer Genome Atlas (TCGA) in which virtually all 30,000 tumors in the archive have been screened with Illumina microarray approaches.…”

Rationale for the development of alternative forms of androgen deprivation therapy

“…Rather, recent ChIP-Seq data have clarified that AR binds to selective AREs because of less stringent sequence requirements for the 3′ hexamer (Sahu, et al 2014). Recent bioinformatics-guided modeling of the ARE motifs derived from similar high-throughput studies indicated that only a minority of the AREs fit perfectly the consensus AGAACAnnnTGTTCT 15-mer motif (Wilson, et al 2016). The majority of AREs corresponded to “imperfect” AREs in which mismatches of 1 or more basepairs from the consensus motif occur.…”

“…Consistent with other reports (see for instance (Massie, et al 2007; Wang, et al 2007)), for a significant number of AR binding sites only one ARE half site that corresponds to a perfect hexamer was present. Increased levels of degeneracy of the ARE have been linked with decreased level of transcriptional output of those sites (Wilson et al 2016). As technology became more sensitive and allowed for higher resolution scanning of the ARE motif and its surrounding sequence, more details could be derived on the genomic regions occupied by AR.…”