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Exploiting native forces to capture chromosome conformation in mammalian cell nuclei.
Molecular Systems Biology 2016 December 10
Mammalian interphase chromosomes fold into a multitude of loops to fit the confines of cell nuclei, and looping is tightly linked to regulated function. Chromosome conformation capture (3C) technology has significantly advanced our understanding of this structure-to-function relationship. However, all 3C-based methods rely on chemical cross-linking to stabilize spatial interactions. This step remains a "black box" as regards the biases it may introduce, and some discrepancies between microscopy and 3C studies have now been reported. To address these concerns, we developed "i3C", a novel approach for capturing spatial interactions without a need for cross-linking. We apply i3C to intact nuclei of living cells and exploit native forces that stabilize chromatin folding. Using different cell types and loci, computational modeling, and a methylation-based orthogonal validation method, "TALE-iD", we show that native interactions resemble cross-linked ones, but display improved signal-to-noise ratios and are more focal on regulatory elements and CTCF sites, while strictly abiding to topologically associating domain restrictions.
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