Elucidating sister chromatid structure by chemical DNA labeling and conformation capture

Chromosomes extensively reorganize during the cell cycle to support regulated gene expression during interphase and to comply with mechanical genome segregation during mitosis. Following chromosome replication in S-phase, the two DNA copies contained in each chromosome co-localize within the same nuclear territory, but later in mitosis appear as separate rod-shaped structures. How sister chromatids fold during interphase and how they resolve to shape mitotic chromosomes has remained unclear. Recently developed chromosome conformation capture methods allow to probe the 3D organization of DNA within cells, yet available technology could not distinguish between sister chromatids. In this project, we developed a new chemical labeling-based methodology to selectively mark sister chromatids for detection by high-throughput DNA sequencing. With this method, termed sister-chromatid-specific Hi-C, we mapped the three-dimensional organization of replicated human chromosomes. We found that a pool of cohesin complexes links sister chromatids at the boundaries of topologically associating domains, whereas another pool of cohesin that forms loops moves sister chromatids apart. Our work provides powerful new technology and insights into fundamental mechanisms shaping the human genome.