Capturing pairwise and multi-way chromosomal conformations using chromosomal walks

Abstract

Chromosomes are folded into highly compacted structures to accommodate physical constraints within nuclei and to regulate access to genomic information. Recently, global mapping of pairwise contacts showed that loops anchoring topological domains (TADs) are highly conserved between cell types and species. Whether pairwise loops synergize to form higher-order structures is still unclear. Here we develop a conformation capture assay to study higher-order organization using chromosomal walks (C-walks) that link multiple genomic loci together into proximity chains in human and mouse cells. This approach captures chromosomal structure at varying scales. Inter-chromosomal contacts constitute only 7-10% of the pairs and are restricted by interfacing TADs. About half of the C-walks stay within one chromosome, and almost half of those are restricted to intra-TAD spaces. C-walks that couple 2-4 TADs indicate stochastic associations between transcriptionally active, early replicating loci. Targeted analysis of thousands of 3-walks anchored at highly expressed genes support pairwise, rather than hub-like, chromosomal topology at active loci. Polycomb-repressed Hox domains are shown by the same approach to enrich for synergistic hubs. Together, the data indicate that chromosomal territories, TADs, and intra-TAD loops are primarily driven by nested, possibly dynamic, pairwise contacts.

Higher-Order Inter-chromosomal Hubs Shape 3D Genome Organization in the Nucleus

“…Although microscopy and specific proximity-ligation methods can also map higher-order interactions (Darrow et al., 2016; Olivares-Chauvet et al, 2016), these are largely restricted to mapping 3-way contacts. In contrast, SPRITE is not restricted in the number of simultaneous DNA contacts.…”

Cell-cycle dynamics of chromosomal organization at single-cell resolution

“…4h, Extended Data Fig. 10e-f), we sampled a randomized ensemble Hi-C map26 and compared contact distributions in the observed and randomized data using the Shaman tool (https://bitbucket.org/tanaylab/shaman). Briefly, we shuffled contacts using a Markov Chain Monte Carlo-like approach, first within each chromosome and then genome-wide, such that the primary factors that define contact distributions are preserved - the marginal coverage and contact distance distribution - but any compartment or TAD structure that may be present is not retained.…”

Nanopore sequencing of DNA concatemers reveals higher-order features of chromatin structure

“…Recent technologies enabling the study of three-way 33 or higher-order chromatin interactions include chromosomal walks (c-walks) 34 , genome architecture mapping (GAM) 35 , split-pool recognition of interactions by tag extension (SPRITE) 36 , ChIA-drop 37 , Tri-C 38 , Tethered multiple 3C 39 , the concatemer ligation assay (COLA) 33 and Multicontact 4C (MC-4C) 40 . Among these assays, only a subset (Tri-C, SPRITE, COLA, GAM) have been shown to generate genome-wide maps in mammals, and all methods but SPRITE suffer from very rare representation of (<1%) of contacts with order >3 36 .…”