Krijger, P. | bioRxiv Science

bioRxiv2021

Implication of the cellular factor CTCF in the regulation of Bovine Leukemia Virus latency and tridimensional chromatin organization

Bovine Leukemia Virus (BLV)-induced tumoral development is a multifactorial phenomenon which remains largely unelucidated. Here, we highlighted the critical role of the cellular CCCTC-binding factor (CTCF) both in the regulation of BLV transcriptional activities and in the deregulation of the tridimensional (3D) chromatin architecture surrounding the BLV integration site. We demonstrated the in vivo recruitment of CTCF to three conserved CTCF binding motifs along the BLV provirus. Next, we showed a critical role for CTCF in delimitating the epigenetic landscape along the BLV provirus as well as to repress the 5Long Terminal Repeat (LTR) promoter activity, thereby contributing to viral latency, while favoring the 3LTR promoter activity. Finally, we demonstrated that BLV integration deregulated host cellular 3D chromatin organization through the formation of abnormal viral/host chromatin loops. Altogether, our results highlight CTCF as a new critical effector of BLV transcriptional regulation and BLV-induced physiopathology.

molecular biology↗

bioRxiv2017

Locus-Specific Enhancer Hubs And Architectural Loop Collisions Uncovered From Single Allele DNA Topologies

Chromatin folding is increasingly recognized as a regulator of genomic processes such as gene activity. Chromosome conformation capture (3C) methods have been developed to unravel genome topology through the analysis of pair-wise chromatin contacts and have identified many genes and regulatory sequences that, in populations of cells, are engaged in multiple DNA interactions. However, pair-wise methods cannot discern whether contacts occur simultaneously or in competition on the individual chromosome. We present a novel 3C method, Multi-Contact 4C (MC-4C), that applies Nanopore sequencing to study multi-way DNA conformations of tens of thousands individual alleles for distinction between cooperative, random and competing interactions. MC-4C can uncover previously missed structures in sub-populations of cells. It reveals unanticipated cooperative clustering between regulatory chromatin loops, anchored by enhancers and gene promoters, and CTCF and cohesin-bound architectural loops. For example, we show that the constituents of the active b-globin super-enhancer cooperatively form an enhancer hub that can host two genes at a time. We also find cooperative interactions between further dispersed regulatory sequences of the active proto-cadherin locus. When applied to CTCF-bound domain boundaries, we find evidence that chromatin loops can collide, a process that is negatively regulated by the cohesin release factor WAPL. Loop collision is further pronounced in WAPL knockout cells, suggestive of a \"cohesin traffic jam\". In summary, single molecule multi-contact analysis methods can reveal how the myriad of regulatory sequences spatially coordinate their actions on individual chromosomes. Insight into these single allele higher-order topological features will facilitate interpreting the consequences of natural and induced genetic variation and help uncovering the mechanisms shaping our genome.

genetics↗