Single-allele chromatin interactions identify regulatory hubs in dynamic compartmentalized domains

Abstract

The promoters of mammalian genes are commonly regulated by multiple distal enhancers, which physically interact within discrete chromatin domains. How such domains form and how the regulatory elements within them interact in single cells is not understood. To address this we developed Tri-C, a new Chromosome Conformation Capture (3C) approach to identify concurrent chromatin interactions at individual alleles. Analysis by Tri-C reveals heterogeneous patterns of single-allele interactions between CTCF boundary elements, indicating that the formation of chromatin domains likely results from a dynamic process. Within these domains, we observe specific higher-order structures involving simultaneous interactions between multiple enhancers and promoters. Such regulatory hubs provide a structural basis for understanding how multiple cis -regulatory elements act together to establish robust regulation of gene expression.

Dynamics of the 4D genome during lineage specification, differentiation and maturation in vivo

“…We focused our analysis on a ~3.3 Mb region containing the well-characterized a-globin genes and their associated regulatory elements. The a-globin genes are regulated by five erythroidspecific enhancer elements (R1-R4 and Rm), which classify as a super-enhancer 20 , and interact with the gene promoters within a TAD flanked by multiple CTCF-binding elements [21][22][23][24] (Supplementary Figure 2). We generated a single-cell RNA-seq dataset 25 , which is the first scRNA-seq dataset to include the full course of in vivo erythroid differentiation through to terminal differentiation in the mouse (Supplementary Figure 5).…”

“…As cells differentiate further, these interactions are strengthened, concomitant with strong upregulation of gene activity. It has recently been shown for the globin loci that gene activation is associated with the formation of higher-order hub-like structures, in which multiple enhancers and promoters form simultaneous, specific interactions 23,31 . Our data suggest that these structures may only be formed in the final stage of differentiation, when chromatin accessibility and interactions between enhancers and promoters are strongest, and may be important to achieve maximal gene expression.…”

CSynth: A Dynamic Modelling and Visualisation Tool for 3D Chromatin Structure

“…In Figure 1 we show data generated from Tri-C data set 18 at the alpha globin region in mouse captured in erythoid cells. Clearly visible is the chromatin looping of the α-globin (mm9, chr11:32,000,000-32,300,000) self-interacting domain (SID).…”

“…As an example of efficacy of CSynth's modelling we use the mouse alpha globin locus ( figure 11). We load matrices generated from TriC data data 18 spanning mouse (mm9) at 4kb resolution. Comparing the resulting model with super resolution microscopy generated using RASER-FISH 30 the chromatin loop or self interacting domain (SID) is clearly visible in the matrix and the model.…”

Enhancers predominantly regulate gene expression in vivo via transcription initiation

“…Changes in RNA expression during cell fate decisions are made via the activation of promoters by enhancers that respond to changes in transcriptional and epigenetic programmes to regulate gene expression in time and space. Many enhancers physically contact target promoters via changes in the three dimensional conformation of the genome (Hay et al 2016;Oudelaar et al 2018;Pennacchio et al 2013;Rowley and Corces 2018;Shlyueva, Stampfel, and Stark 2014). This may increase the concentration of transcription factors, co-factors, and Pol II at the promoters of their target genes Cho et al 2018;Cramer 2019;Heinz et al 2015;J.…”