CAPTURE-3C-seq analysis of cis-element-mediated long-range DNA interactions

Cis-regulatory elements (CREs) are commonly recognized by correlative chromatin features, yet the molecular composition of the vast majority of CREs in chromatin remains unknown. Here we describe a CRISPR affinity purification in situ of regulatory elements (CAPTURE) approach to unbiasedly identify locus-specific chromatin-regulating protein complexes and long-range DNA interactions. Using an in vivo biotinylated endonuclease-deficient Cas9 protein and sequence-specific guide RNAs, we show high-resolution and selective isolation of chromatin interactions at a single copy genomic locus. Purification of human telomeres using CAPTURE identifies known and new telomeric factors. In situ capture of individual constituents of the enhancer cluster controlling human ß-globin genes establishes evidence for composition-based hierarchical organization of enhancer structure. Furthermore, unbiased analysis of chromatin interactions at disease-associated cis-elements and developmentally controlled super-enhancers reveals spatial features causally regulate gene transcription. Thus, comprehensive analysis of locus-specific regulatory composition provides mechanistic insight into genome structure and function in development and disease. Overall design: We combined the chromatin interaction assays (Chromatin Conformation Capture or 3C) with the high affinity dCas9 purification approach to unbiasedly identify single genomic locus-associated long-range interactions (CAPTURE-3C-seq).