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Cohesin reshapes replication fork contacts to aid fork slowing and reversal
Nature
(2026) Cite this article
DNA replication forks can be challenged by cancer chemotherapeutic treatments, leading to accumulation of single-stranded DNA and slowdown of DNA synthesis. The marked plasticity of replication forks under replication stress ensures fork stability, damage tolerance and complete genome duplication1. Initiation and progression of replication forks occur in a three-dimensionally organized genome. DNA loop extrusion by the cohesin complex organizes the genome2 and regulates the initiation and positioning of DNA replication origins3,4. Although transient interaction of sister forks was recently reported during unperturbed replication5, the functional relevance of fork contacts during replication stress and the role of cohesin in this context remain unknown. Here we show that cohesin-mediated loop extrusion rearranges nascent DNA contacts at stressed replication forks to promote genome stability. Using auxin-inducible degron6, separation-of-function mutants7,8,9 and a newly developed Micro-C-based technique to capture chromatin contacts at nascent DNA (Repli-C), we found that loop-extruding cohesin accumulates at stalled replication forks, limiting sister-fork coupling in favour of inter-replicon contacts. This process promotes active fork slowing and reversal by preventing PRIMPOL action on single-stranded DNA1. These findings show that the replication stress response is not merely an accumulation of individual regulatory events, but is topologically integrated across the genome through cohesin loop extrusion. While providing a new function for loop-extruding cohesin, our results indicate the potential impact on cancer therapy of frequent cohesin mutations in tumours10.
Replication of organized genomes is spatiotemporally regulated, in which topologically associating domains (TADs) containing marks of open chromatin and active transcription replicate earlier than compact, transcriptionally repressed TADs11,12. Initiation of DNA replication takes place at DNA replication origins that spread along the genome13,14, which are enriched in initiation zones, defined by population studies15. Recent replication-specific Hi-C analyses revealed nascent DNA contacts within the same initiation zone, possibly representing sister forks of the same replicon and/or converging forks from neighbouring replicons, which are physically coupled during DNA synthesis5. However, how nascent DNA organization is affected by conditions interfering with fork progression (that is, replication stress (RS)) remains unknown. RS triggers changes in the composition of the replication apparatus and in the architecture of replication forks. This ‘fork plasticity’ assists cells tolerating DNA damage and replication interference1,11. One such mechanism is fork reversal, the active and reversible remodelling of the replication fork into a four-way junction through annealing of the two nascent DNA strands. This requires RAD51 and the DNA translocases SMARCAL1, HLTF and ZRANB3. An alternative mechanism sustaining DNA synthesis under stress is repriming by the DNA primase and polymerase PRIMPOL to promote discontinuous DNA synthesis1. How these mechanisms are used locally and globally remains unknown and may reflect chromatin architecture and the accessibility of nascent DNA to different stress-tolerance factors.
The cohesin complex (SMC1, SMC3, RAD21 and either STAG1 or STAG2) organizes the genome into TADs and chromatin loops because of its ability to extrude DNA16,17,18. This function requires its association with the cohesin chromatin loader NIPBL–MAU2 (refs. 2,19). Loop-extruding cohesin is stopped at certain locations by protein barriers, such as CTCF, or the MCM complex belonging to the replicative CMG helicase (CDC45–MCM–GINS)20,21. Cohesin also interacts with replicative pre-initiation complexes and aids localizing replication origins and initiation zones at early replicating domains3,4. During DNA repl