Abstract
Using a chromatin immunoprecipitation-paired end diTag cloning and sequencing strategy, we mapped estrogen receptor α (ERα) binding sites in MCF-7 breast cancer cells. We identified 1,234 high confidence binding clusters of which 94% are projected to be bona fide ERα binding regions. Only 5% of the mapped estrogen receptor binding sites are located within 5 kb upstream of the transcriptional start sites of adjacent genes, regions containing the proximal promoters, whereas vast majority of the sites are mapped to intronic or distal locations (>5 kb from 5′ and 3′ ends of adjacent transcript), suggesting transcriptional regulatory mechanisms over significant physical distances. Of all the identified sites, 71% harbored putative full estrogen response elements (EREs), 25% bore ERE half sites, and only 4% had no recognizable ERE sequences. Genes in the vicinity of ERα binding sites were enriched for regulation by estradiol in MCF-7 cells, and their expression profiles in patient samples segregate ERα-positive from ERα-negative breast tumors. The expression dynamics of the genes adjacent to ERα binding sites suggest a direct induction of gene expression through binding to ERE-like sequences, whereas transcriptional repression by ERα appears to be through indirect mechanisms. Our analysis also indicates a number of candidate transcription factor binding sites adjacent to occupied EREs at frequencies much greater than by chance, including the previously reported FOXA1 sites, and demonstrate the potential involvement of one such putative adjacent factor, Sp1, in the global regulation of ERα target genes. Unexpectedly, we found that only 22%–24% of the bona fide human ERα binding sites were overlapping conserved regions in whole genome vertebrate alignments, which suggest limited conservation of functional binding sites. Taken together, this genome-scale analysis suggests complex but definable rules governing ERα binding and gene regulation.
Highlights
Cellular transcriptomes are dictated by complex interactions between signal transduction pathways, general and specific transcription factors, chromatin remodeling proteins, and the RNA polymerase complexes
To obtain a genome-wide view of estrogen receptor a (ERa) binding sites, we applied chromatin immunoprecipitation coupled with a cloning and sequencing strategy using chromatin immunoprecipitation pair end-tagging technology to map ERa binding sites in MCF-7 human breast cancer cells
We identified 1,234 high quality ERa binding sites in the human genome and demonstrated that the binding sites are frequently adjacent to genes significantly associated with breast cancer disease status and outcome
Summary
Cellular transcriptomes are dictated by complex interactions between signal transduction pathways, general and specific transcription factors, chromatin remodeling proteins, and the RNA polymerase complexes. Precise transcriptional responses are achieved in part by targeting transcription factor complexes to the cis-regulatory regions of target genes via specific binding site sequences The importance of these binding site sequences is reflected in the conservation of sequence motifs in coregulated genes and through evolution. The mechanisms of ER binding site specificity, are not clear since these binding site sequence motifs are ubiquitous in the genome, and there is no discernable difference between functional and nonfunctional sites by computational modeling approaches. This ambiguity is likely due to a lack of systemic
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call