Cohesin-dependent chromatin structure at high resolution.

Genome-wide studies to address the three-dimensional structure of different genomes identified a general and conserved organization in topological domains. Recently, we demonstrated that the cohesin complex has an important role in the organization of the topological domains. By using 3C-sequencing (3C-seq) to investigate local changes in the chromatin structure and Hi-C to study genome-wide changes in chromosomal interactions we showed that the depletion of cohesin by the proteolytic cleavage of its RAD21 subunit leads to reduction of chromatin interactions, predominantly within topological domains. However, the limitation to use just a restricted number of DNA fragments as viewpoint for the 3C-seq and the resolution of Hi-C (40kb) are not sufficient to yield information about how the fiber is folded within these domains and which are the detailed effects of the cohesin depletion on the interacting areas. To address this question we used a high resolution approach called “Targeted Chromatin Capture (T2C)”. T2C provides information of the spatial organization of selected loci at singlerestriction fragment resolution (2 to 6 kbp). We studied a genomic region of 2.1 Mb on the human chromosome 11 comprising the imprinting loci IGF2/H19 and KCNQ1 to unravel the details. Here we will present our latest results.