Far from being a static archive of information, the genome is composed of dynamic molecules that fold, replicate, and respond to the chemistry of life. These processes, along with external stresses such as radiation or reactive molecules, expose DNA to wear and tear, creating damage that the cell must detect and repair to keep the genome intact. Among the most dangerous lesions are DNA double-strand breaks—cuts through the double helix that can lead to harmful rearrangements of chromosomes, mutations, or cell death if not repaired swiftly and precisely.

The most accurate way to fix such breaks is through homology-directed repair, a pathway in which the cell restores the missing information by copying it from an identical sequence, usually the sister chromatid. But finding this matching sequence within a genome that contains billions of DNA letters is a “needle-in-a-haystack” search.

Although scientists had identified many of the molecular tools involved in this repair pathway, how these tools find the right place to work inside the crowded nucleus was still unknown. Cohesin—a ring-shaped protein that holds sister chromatids together and folds DNA into loops—was known to be essential for the process, yet its role in guiding the homology search had not been resolved.

Now, researchers from the lab of Daniel Gerlich at IMBA have discovered how cohesin guides this homology search. Their study, published in the journal Science, reveals that cohesin operates in two coordinated modes during DNA repair: one remodels the surrounding chromatin to enlarge loops that define where the search takes place;  the other keeps the broken DNA physically linked to its undamaged sister copy. Together, these mechanisms transform a daunting genome-wide quest into a focused and efficient search within the nucleus.

When DNA breaks, cohesin leads the way

To discover how broken DNA finds its match within the crowded nucleus, the team combined precise induction of DNA breaks with their newly developed technique sister-pore-C. This method pairs a clever DNA-labeling strategy with sequencing to distinguish physical interactions within a single chromatid from those between sister chromatids—something standard sequencing could not resolve. Sister-pore-C builds on a related technique previously developed in the Gerlich lab, but provides much higher-resolution maps. This improvement makes it possible to track how sister chromatids interact before and after a break, and to reveal in unprecedented detail how the structure of chromosomes reorganizes around damaged DNA in human cells.

“Until now, we simply lacked a way to see how sister chromatids interact inside living cells,” says Federico Teloni, postdoc in the Gerlich lab and first author of the study. “Because the two DNA molecules are identical, traditional sequencing cannot tell them apart. With this new approach, we can finally distinguish the sisters and watch how a broken DNA end connects with its sister to locate the matching sequence needed for repair.”

Their results reveal a striking choreography. After a DNA break occurs, one group of cohesin complexes begins to remodel the surrounding chromatin by extruding loops across a broad, megabase-scale region—essentially outlining where the search for the matching sequence should take place. At the same time, another set of cohesin molecules forms a clamp directly at the broken DNA end. Instead of drifting freely through the nucleus, the damaged DNA remains anchored to its sister chromatid, where the correct sequence resides. As loop extrusion continues, this anchored end is carried along the sister chromatid in a directed, linear manner rather than randomly exploring the whole nuclear space.

The researchers also showed that both cohesin functions are indispensable. When loop extrusion is disrupted, the search becomes confined and inefficient, slowing the repair process. When the stabilizing hold between the broken DNA and its sister copy is lost, the repair cannot proceed at all. By combining broad-scale looping with a local anchoring mechanism, cohesin transforms what would otherwise be a random, three-dimensional hunt into a directed search along the sister chromatid.

This work reveals that chromosome architecture is not a passive scaffold but an active determinant of DNA repair. Cohesin not only holds sister chromatids together—it shapes how the homology search unfolds in space and time.

“This study gives us a new insight into how cells handle DNA molecules that are tens of thousands of times longer than the cell itself,” says Daniel Gerlich, senior scientist at IMBA. “Understanding how the three-dimensional organization of the genome contributes to DNA repair could eventually help improve genome engineering or other technologies that rely on precise DNA repair.”

“This project simply wouldn’t have been possible without the collaborative environment at the Vienna BioCenter,” Gerlich adds. “It’s a perfect example of what happens when ideas move freely between groups — you can create something no single field could have invented alone.”