Research Article

Single-molecule regulatory architectures captured by chromatin fiber sequencing

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Science  26 Jun 2020:
Vol. 368, Issue 6498, pp. 1449-1454
DOI: 10.1126/science.aaz1646

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Primary architecture of chromatin fibers

The organization of chromosomal DNA, including the positioning of nucleosomes and nucleosome-free regions harboring regulatory proteins along single chromatin fibers, is fundamental to genome function. However, most sequencing methods cannot elucidate this organization at the nucleotide level. Stergachis et al. present an approach, Fiber-seq, that maps chromatin fibers onto the underlying DNA template using methyltransferases to create a kind of stencil in fly and human cells. This method identifies chromatin structure at nearly a single-molecule level and can monitor the position of nucleosomes. Using Fiber-seq, the authors identify how regulatory DNA activation is related to nucleosome positioning and DNA variation.

Science, this issue p. 1449


Gene regulation is chiefly determined at the level of individual linear chromatin molecules, yet our current understanding of cis-regulatory architectures derives from fragmented sampling of large numbers of disparate molecules. We developed an approach for precisely stenciling the structure of individual chromatin fibers onto their composite DNA templates using nonspecific DNA N6-adenine methyltransferases. Single-molecule long-read sequencing of chromatin stencils enabled nucleotide-resolution readout of the primary architecture of multikilobase chromatin fibers (Fiber-seq). Fiber-seq exposed widespread plasticity in the linear organization of individual chromatin fibers and illuminated principles guiding regulatory DNA actuation, the coordinated actuation of neighboring regulatory elements, single-molecule nucleosome positioning, and single-molecule transcription factor occupancy. Our approach and results open new vistas on the primary architecture of gene regulation.

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