How do the chromosomal DNA molecules that encode the genome, that measure almost two meters, fit into cells that are a fraction of the size? The answer lies in the spatial organization of chromosomal DNA which enables regulated topological interactions between distant parts, thereby supporting proper expression, maintenance, and transport of the genome across cell generations. The three-dimensional organization of the genome also supports regulated recombination, DNA repair, and chromosome segregation during mitosis.

For some DNA repair mechanisms to occur, the two DNA molecules of sister chromatids need to come close together at the exact same genomic position. How the two DNA molecules are organized relative to each other to support this important repair pathway, however, has remained unclear.

Chromosome conformation capture (Hi-C) analysis has previously revealed complex internal chromosomal structures in vertebrate cells, but the identical sequence of sister chromatids has made it difficult to determine how they interact in replicated chromosomes.

Now, a team from the Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), in Vienna, describe sister-chromatid-sensitive Hi-C (scsHi-C), which is based on the labeling of nascent DNA with 4-thio-thymidine and nucleoside conversion chemistry. In doing so, they have created the first high resolution map of contact points between replicated chromosomes.

The work is published in an article in Nature titled, “Conformation of sister chromatids in the replicated human genome.

The team around Daniel Gerlich, PhD, senior group leader at the IMBA, developed a method to create the first high resolution map of contact points between replicated chromosomes. “Current methods to map the folding of DNA have a serious blind spot: They are not able to distinguish identical copies of DNA molecules. Our approach to solve this was to label DNA copies in a way such that we can discriminate them by DNA sequencing,” explained Michael Mitter, a doctoral student in Gerlich’s lab. Using this approach, the researchers were able to create the first high resolution map of contact points between replicated chromosomes.

The authors write that, “Genome-wide conformation maps of human chromosomes reveal that sister-chromatid pairs interact most frequently at the boundaries of topologically associating domains (TADs).” They continued: “Continuous loading of a dynamic cohesin pool separates sister-chromatid pairs inside TADs and is required to focus sister-chromatid contacts at TAD boundaries.” The authors identified a subset of TADs that are overall highly paired and are characterized by facultative heterochromatin and insulated topological domains that form separately within individual sister chromatids.

“With this new method, we can now study the molecular machinery regulating the conformation of sister chromatids, which will provide insights into the mechanics underlying the repair of DNA breaks and the formation of rod-shaped chromosomes in dividing cells, which is required for proper transport the genome to cell progeny,” said Gerlich about the project, which is financed by the Vienna Science and Technology Fund (WWTF) and was a fruitful collaboration of several research groups at the Vienna BioCenter, including the Ameres and Goloborodko labs at IMBA, and the Peters lab at the neighboring Institute of Molecular Pathology (IMP).

The authors explained that the rich pattern of sister-chromatid topologies and their scsHi-C technology will “make it possible to investigate how physical interactions between identical DNA molecules contribute to DNA repair, gene expression, chromosome segregation, and potentially other biological processes.”

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