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Alberto Marin @albertomarin.bsky.social · 12/06/2026

We found that loops on the broken chromatid promote the initial encounter between the break and the sister chromatid by steering the break toward TAD boundaries. Loops on the sister chromatid drive the subsequent 1D scanning that is the rate-limiting step for homology identification. 8/n

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

We discovered that instead of searching randomly, cells use an active 1D scanning process: the repair machinery leverages a looping protein called cohesin to ‘slide’ the break along the DNA and find the matching sequence. 6/n

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

Simply relying on random 3D diffusion – letting the broken DNA wander through the nucleus – would be inefficient. Even for a 1 Mb region, the broken DNA would take far too long to find its matching sequence by chance alone. 5/n

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

Now, after DNA replicates, sister chromatids are held together approximately every 1 Mb so the search is confined to ~1 M nucleotides. But finding the right match is still a huge challenge – especially if the sisters aren’t perfectly aligned. 4/n

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

When DNA breaks, cells often repair it through a process called homologous recombination, in which a matching (replicated) copy of the broken sequence is used as a repair template. 2/n

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

Link to the publication below! 8/n www.science.org/doi/10.1126/...

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

Thus, chromatin loops don’t just organize the genome to control gene expression – they also protect its integrity by helping a broken DNA find its matching sequence for repair! 7/n

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

Homologous recombination is key for protecting the genome, but it’s also challenging because the broken DNA must find its matching copy within billions of nucleotides. How can a cell achieve this? This is know as the “homology search” problem. 3/n

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Alberto Marin @albertomarin.bsky.social · 04/12/2025

When DNA breaks, cells often repair it through a process called homologous recombination, in which a matching (replicated) copy of the broken sequence is used as a repair template. 2/n

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

12/n Our model, in a nutshell: cohesin drives homology search via 1D scanning. During HR, a RAD51 filament locally scans the sister chromatid, but this search could be unproductive (e.g., because the donor is far). Cohesin loops would then facilitate long-range scanning to help find a donor!

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

11/n Does loop-extruding cohesin (Nipbl) regulates HR? In the short-range HR-GFP reporter, Nipbl depletion mildly reduces HR (45%). But in the long-range reporters, Nipbl depletion substantially reduces HR (75%). Thus, Nipbl is critical for HR when long searches are required.

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

10/n We noticed that the RAD51 domain is constrained by TAD boundaries, suggesting a role for cohesin in mediating the homology search. Indeed, acute degradation of the cohesin unloader WAPL, known to yield elongated loops, resulted in a broader search.

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

9/n So, we generated monoclonal mES cell lines with the donor at +441 kb and +563 kb from the break. When we induced the DSB…we found a beautiful RAD51 peak at the exact location of the donor! The RAD51 chromatin domain is thus capturing the homology search.

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

/n But why does RAD51 spread to such long distances (~0.5 Mb) from the break? End resection was constrained to ~ 5 kb, so the broad RAD51 domain does not come from the RAD51-ssDNA filament. We hypothesized that the RAD51 domain reflects the homology search. We tested this.

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

6/n Motivated by this finding, we performed a second time-course experiment, where we looked at the dynamics of RAD51 recruitment, the core HR factor. RAD51 dynamics mimic break-anchored loop formation, suggesting a causal relationship between loops and HR.

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

5/n To answer this, we induced DSBs in cell-cycle-synchronized populations. Cells in late S/G2 phases, which are HR-proficient, showed break-anchored loops. But cells synchronized in G1, which are HR-deficient, did not show the loops. Break-anchored loops are HR events!

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

4/n Time-course Hi-C, enabled by our light-activated very fast CRISPR, showed that break-anchored loops are late repair events. Loops form after γH2AX domain, ruling out previous models where break-anchored loops propagate γH2AX at early stages. What is the role of such loops?

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

3/n Using our multi-target CRISPR system – which induces hundreds of DNA double-strand breaks (DSB) on demand – and Hi-C, we found that Cas9 breaks act as anchors for chromatin loops. Cohesin is recruited to the breaks and is required for break-anchored loop formation.

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Alberto Marin @albertomarin.bsky.social · 13/02/2025

2/n The homologous recombination (HR) pathway repairs DNA lesions by copying intact sequence information from a DNA template… but how does a damaged DNA find a correct DNA template in the 3D genome? We provide some answers to this Q, known as the homology search problem.

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