Article | Published:

Biochemical reconstitution of topological DNA binding by the cohesin ring

Nature volume 505, pages 367371 (16 January 2014) | Download Citation

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Abstract

Cohesion between sister chromatids, mediated by the chromosomal cohesin complex, is a prerequisite for faithful chromosome segregation in mitosis. Cohesin also has vital roles in DNA repair and transcriptional regulation. The ring-shaped cohesin complex is thought to encircle sister DNA strands, but its molecular mechanism of action is poorly understood and the biochemical reconstitution of cohesin activity in vitro has remained an unattained goal. Here we reconstitute cohesin loading onto DNA using purified fission yeast cohesin and its loader complex, Mis4Scc2–Ssl3Scc4 (Schizosaccharomyces pombe gene names appear throughout with their more commonly known Saccharomyces cerevisiae counterparts added in superscript). Incubation of cohesin with DNA leads to spontaneous topological loading, but this remains inefficient. The loader contacts cohesin at multiple sites around the ring circumference, including the hitherto enigmatic Psc3Scc3 subunit, and stimulates cohesin’s ATPase, resulting in efficient topological loading. The in vitro reconstitution of cohesin loading onto DNA provides mechanistic insight into the initial steps of the establishment of sister chromatid cohesion and other chromosomal processes mediated by cohesin.

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Acknowledgements

We are grateful to N. O’Reilly for peptide synthesis, A. Alidoust and N. Patel for fermentation and J. Hurwitz, T. Toda and members of the Chromosome Segregation Laboratory for discussion and comments on the manuscript. This work was supported by the European Research Council. Y.M. was supported by the Japanese Society for the Promotion of Science (JSPS).

Author information

Affiliations

  1. Chromosome Segregation Laboratory, Cancer Research UK London Research Institute, 44 Lincoln’s Inn Fields, London WC2A 3LY, UK

    • Yasuto Murayama
    •  & Frank Uhlmann

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Contributions

Y.M. designed the study, performed all the experiments, analysed data and wrote the manuscript. F.U. designed and supervised the study and wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Frank Uhlmann.

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https://doi.org/10.1038/nature12867

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