CTCF modulates allele-specific sub-TAD organization and imprinted gene activity at the mouse Dlk1-Dio3 and Igf2-H19 domains

Abstract

BackgroundGenomic imprinting is essential for mammalian development and provides a unique paradigm to explore intra-cellular differences in chromatin configuration. So far, the detailed allele-specific chromatin organization of imprinted gene domains has mostly been lacking. Here, we explored the chromatin structure of the two conserved imprinted domains controlled by paternal DNA methylation imprints—the Igf2-H19 and Dlk1-Dio3 domains—and assessed the involvement of the insulator protein CTCF in mouse cells.ResultsBoth imprinted domains are located within overarching topologically associating domains (TADs) that are similar on both parental chromosomes. At each domain, a single differentially methylated region is bound by CTCF on the maternal chromosome only, in addition to multiple instances of bi-allelic CTCF binding. Combinations of allelic 4C-seq and DNA-FISH revealed that bi-allelic CTCF binding alone, on the paternal chromosome, correlates with a first level of sub-TAD structure. On the maternal chromosome, additional CTCF binding at the differentially methylated region adds a further layer of sub-TAD organization, which essentially hijacks the existing paternal-specific sub-TAD organization. Perturbation of maternal-specific CTCF binding site at the Dlk1-Dio3 locus, using genome editing, results in perturbed sub-TAD organization and bi-allelic Dlk1 activation during differentiation.ConclusionsMaternal allele-specific CTCF binding at the imprinted Igf2-H19 and the Dlk1-Dio3 domains adds an additional layer of sub-TAD organization, on top of an existing three-dimensional configuration and prior to imprinted activation of protein-coding genes. We speculate that this allele-specific sub-TAD organization provides an instructive or permissive context for imprinted gene activation during development.

ASHIC: Hierarchical Bayesian modeling of diploid chromatin contacts and structures

“…Specifically, these CTCF clusters interacted preferentially with H19 on the maternal allele and with Igf2 on the paternal allele. These observations are consistent with the previous 4C-seq results in mouse ESCs [33].…”

“…For the quantitative comparison, we calculated the pairwise Euclidean distances of mHIDAD, H19, and Igf2 on the maternal and paternal structures predicted by ASHIC-ZIPM from 20 random initializations. As shown in Figure 8C, the distance between mHIDAD and H19 was significantly smaller on the maternal structure than that on the paternal structure (p-value = 4.43 × 10 −5 , one-sided Wilcoxon paired signed-rank test), which is consistent with the previous DNA-FISH data [33]. In contrast, the distance between mHIDAD and Igf2 was significantly larger on maternal allele (p-value = 4.43 × 10 −5 , one-sided Wilcoxon paired signed-rank test), which is consistent with the observation of paternal-specific HIDAD-IGF2 loop in human GM12878 cells [6].…”

“…This sub-TAD formation creates an insulation between H19 and Igf2 that likely prevents the activation of Igf2 on the maternal allele. Furthermore, the diploid Hi-C maps produced by ASHIC-ZIPM offer an informative view of the (sub-)TAD organizations on the imprinting region, whereas the previous 4C-seq study [33] were restricted to only few anchor regions.…”

“…Unlike TADs that are mostly invariant between cell types, sub-TADs are more variable and play a pivotal role in mediating cell-type-specific gene regulation [39,40]. Based on the presence of monoallelic CTCF bindings at H19-DMR, Llères et al [33] proposed a novel parental-specific sub-TAD model for the regulation of imprinting at H19/Igf2 locus. Supported by allelic 4C-seq and DNA-FISH data, they speculated that several bi-allelic CTCF binding sites form a first layer of TAD on both alleles.…”

Haldane’s rule in the placenta: sex-biased misregulation of theKcnq1imprinting cluster in hybrid mice

“…Smarca1 encodes a chromatin remodeling enzyme that 551 suppresses cell proliferation and attenuates Wnt-signaling (Eckey et al 2012), processes crucial 552 for embryogenesis and placental development (Parr et al 2001;Logan and Nusse 2004). 553 Members of the Smarca gene family promote insulator protein CTCF binding with downstream 554 effects on CTCF-regulated genes (Wiechens et al 2016), which include several imprinted gene 555 clusters (Lléres et al 2019). Given that Smarca1 was transgressively overexpressed in hybrid 556 male placenta, the potential contribution of this gene to the difference between hybrid male and 557 female phenotypes is worthy of further study.…”