Genomes are organized into high-level 3-dimensional structures, and DNA elements separated by long genomic distances could functionally interact. Many transcription factors bind to regulatory DNA elements distant from gene promoters. While distal binding sites have been shown to regulate transcription by long-range chromatin interactions at a few loci, chromatin interactions and their impact on transcription regulation have not been investigated in a genome-wide manner. Therefore, we developed Chromatin Interaction Analysis by Paired-End Tag sequencing (ChIA-PET) for de novo detection of global chromatin interactions, and comprehensively mapped the chromatin interaction network bound by oestrogen receptor α (ERα) in the human genome. We found that most high-confidence remote ERα binding sites are anchored at gene promoters through long-range chromatin interactions, suggesting that ERα functions by extensive chromatin looping to bring genes together for coordinated transcriptional regulation. We propose that chromatin interactions constitute a primary mechanism for regulating transcription in mammalian genomes.

Physiological Wnt signaling is required for the maintenance of the crypt progenitor phenotype and controls the proliferation/differentiation switch in the adult, self-renewing intestinal epithelium (33). A constitutively active Tcf/␤-catenin transcription complex, resulting from mutations in adenomatous polyposis coli (APC), Axin, or ␤-catenin, is the primary transforming factor in colorectal cancer (CRC) (25,26,32); aberrant Tcf/␤-catenin activity results in a transcriptional profile in CRC cells similar to that which is physiologically driven by Tcf/␤-catenin in the crypt stem/progenitor cells of the intestine (49). Through candidate gene approaches and microarray technology, a large number of genes have been uncovered whose expression levels are altered upon abrogation or activation of the Wnt pathway (for references, see http://www .stanford.edu/ϳrnusse/pathways/targets.html). It remains unclear whether the affected genes are direct or indirect targets of the Tcf/␤-catenin transcription factor complex. cis-regulatory elements directly bound by Tcf have been identified for only a few candidate genes. Such studies have been mostly limited to regulatory regions close to the transcription start site (TSS) of candidate genes (e.g., see reference 17). A comprehensive identification of regulatory elements is essential for a more complete understanding of the transcriptional repertoire driven by the Wnt pathway and the elucidation of the molecular mechanisms by which Tcf and ␤-catenin control the transcription of their target genes.A recent approach taken to achieve such goals is chromatin immunoprecipitation (ChIP)-coupled DNA microarray analysis (ChIP-on-chip), which couples the immunoprecipitation of chromatin-bound transcription factors with the identification of the bound DNA sequences through hybridization on DNA microarrays (35). This approach has been used to generate, among others, a comprehensive map of active, preinitiation complex-bound promoters in human fibroblast cells (24). Microarrays covering the nonrepetitive sequence of chromosomes 21 and 22 have allowed the study of histone H3 methylation and acetylation patterns in human hepatoma cells (5) and estrogen receptor binding sites in breast cancer cells (8). The latter study revealed selective binding of estrogen receptor (ER) to a limited number of sites, most of which were distant from the TSSs of ER-regulated genes (8). Similar conclusions were put forth by work examining the in vivo binding of transcription factors Sp1, c-Myc, and p53 along chromosomes 21 and 22: most binding sites identified do not correspond to the proximal promoters of protein-coding genes but rather lie within or immediately 3Ј to well-characterized genes or are significantly correlated with noncoding RNAs (10). Collectively these studies point to the necessity of interrogating entire genomes for the comprehensive determination of in vivo-occupied binding sites (9,23,52,54).In the present work, we used a combination of ChIP and location analysis with genome-wide tiling arrays...

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