Bioinformatics Methods Analysize CTCF Function In Genome Topology And Enhancer/Promoter Interaction

CTCF and the associated cohesin complex play a central role in insulator function and higher-order chromatin organization of mammalian genomes.Recent studies identified a correlation between the orientation of CTCF-binding sites(CBSs)and chromatin interactions.To test the functional significance of this observation,we combined CRISPR/Cas9-based genomic-DNA-fragment editing with chromosome conformation capture sequencing to show that the location and relative orientations of CBSs determine the specificity of long-range chromatin looping in mammalian genomes,using protocadherin(Pcdh)and ?-globin as model genes.Inversion of CBS elements within the Pcdh enhancer reconfigures the topology of chromatin loops between the distal enhancer and target promoters and alters gene-expression patterns.We use the published and ENCODE ChIP-seq and ChIA-PET data,and analysis the genome-wide chromosome interactions relate with CTCF/cohesin binding to the CBS sites and relate CCDs(CTCF/Cohesin-mediated Chromatin Domain)boundary CBS pairs,and we found that most those interactions are Forward-Reverse orientations.We also found that more than 90% of CCDs boundaries CBS pairs are Reverse-Forward.Thus,although enhancers can function in an orientation-independent manner in reporter assays,in the native chromosome context,the orientation of at least some enhancers carrying CBSs can determine both the architecture of topological chromatin domains and enhancer/promoter specificity.These findings reveal how 3D chromosome architecture encoded by linear genome sequences.