Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Recruitment of Xenopus Scc2 and cohesin to chromatin requires the pre-replication complex

Nature Cell Biology volume 6, pages 991–996 (2004)Cite this article

Abstract

Cohesin is a multi-subunit, ring-shaped protein complex that holds sister chromatids together from the time of their synthesis in S phase until they are segregated in anaphase1,2,3,4,5,6,7. In yeast, the loading of cohesin onto chromosomes requires the Scc2 protein8,9,10. In vertebrates, cohesins first bind to chromosomes as cells exit mitosis, but the mechanism is unknown3,11,12. Concurrent with cohesin binding, pre-replication complexes (pre-RCs) are assembled at origins of DNA replication through the sequential loading of the initiation factors ORC, Cdc6, Cdt1 and MCM2-7 (the 'licensing' reaction)13. In S phase, the protein kinase Cdk2 activates pre-RCs, causing origin unwinding and DNA replication. Here, we use Xenopus egg extracts to show that the recruitment of cohesins to chromosomes requires fully licensed chromatin and is dependent on ORC, Cdc6, Cdt1 and MCM2-7, but is independent of Cdk2. We further show that Xenopus Scc2 is required for cohesin loading and that binding of XScc2 to chromatin is MCM2-7 dependent. Our results define a novel pre-RC-dependent pathway for cohesin recruitment to chromosomes in a vertebrate model system.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Chromatin loading of x-cohesin is inhibited by geminin.
Figure 2: x-cohesin loading is dependent on pre-RC formation.
Figure 3: x-cohesin loading is independent of Cdk2 and origin unwinding.
Figure 4: XScc2, which is required for cohesin loading, binds to chromatin in a pre-RC-dependent manner.

Similar content being viewed by others

Accession codes

Accessions

GenBank/EMBL/DDBJ

References

  1. Michaelis, C., Ciosk, R. & Nasmyth, K. Cohesins: chromosomal proteins that prevent premature separation of sister chromatids. Cell 91, 35–45 (1997).

    Article  CAS  Google Scholar 

  2. Uhlmann, F. & Nasmyth, K. Cohesion between sister chromatids must be established during DNA replication. Curr. Biol. 8, 1095–1101 (1998).

    Article  CAS  Google Scholar 

  3. Losada, A., Hirano, M. & Hirano, T. Identification of Xenopus SMC protein complexes required for sister chromatid cohesion. Genes Dev. 12, 1986–1997 (1998).

    Article  CAS  Google Scholar 

  4. Guacci, V., Koshland, D. & Strunnikov, A. A direct link between sister chromatid cohesion and chromosome condensation revealed through the analysis of MCD1 in S. cerevisiae. Cell 91, 47–57 (1997).

    Article  CAS  Google Scholar 

  5. Uhlmann, F., Lottspeich, F. & Nasmyth, K. Sister-chromatid separation at anaphase onset is promoted by cleavage of the cohesin subunit Scc1. Nature 400, 37–42 (1999).

    Article  CAS  Google Scholar 

  6. Gruber, S., Haering, C.H. & Nasmyth, K. Chromosomal cohesin forms a ring. Cell 112, 765–777 (2003).

    Article  CAS  Google Scholar 

  7. Nasmyth, K. Disseminating the genome: joining, resolving, and separating sister chromatids during mitosis and meiosis. Annu. Rev. Genet. 35, 673–745 (2001).

    Article  CAS  Google Scholar 

  8. Toth, A. et al. Yeast cohesin complex requires a conserved protein, Eco1p(Ctf7), to establish cohesion between sister chromatids during DNA replication. Genes Dev. 13, 320–333 (1999).

    Article  CAS  Google Scholar 

  9. Tomonaga, T. et al. Characterization of fission yeast cohesin: essential anaphase proteolysis of Rad21 phosphorylated in the S phase. Genes Dev. 14, 2757–2770 (2000).

    Article  CAS  Google Scholar 

  10. Ciosk, R. et al. Cohesin's binding to chromosomes depends on a separate complex consisting of Scc2 and Scc4 proteins. Mol. Cell 5, 243–254 (2000).

    Article  CAS  Google Scholar 

  11. Darwiche, N., Freeman, L.A. & Strunnikov, A. Characterization of the components of the putative mammalian sister chromatid cohesion complex. Gene 233, 39–47 (1999).

    Article  CAS  Google Scholar 

  12. Sumara, I., Vorlaufer, E., Gieffers, C., Peters, B.H. & Peters, J.M. Characterization of vertebrate cohesin complexes and their regulation in prophase. J. Cell Biol. 151, 749–762 (2000).

    Article  CAS  Google Scholar 

  13. Bell, S.P. & Dutta, A. DNA replication in eukaryotic cells. Annu. Rev. Biochem. 71, 333–374 (2002).

    Article  CAS  Google Scholar 

  14. Walter, J., Sun, L. & Newport, J. Regulated chromosomal DNA replication in the absence of a nucleus. Mol. Cell 1, 519–529 (1998).

    Article  CAS  Google Scholar 

  15. Hodgson, B., Li, A., Tada, S. & Blow, J.J. Geminin becomes activated as an inhibitor of Cdt1/RLF-B following nuclear import. Curr. Biol. 12, 678–683 (2002).

    Article  CAS  Google Scholar 

  16. Tada, S., Li, A., Maiorano, D., Mechali, M. & Blow, J.J. Repression of origin assembly in metaphase depends on inhibition of RLF-B/Cdt1 by geminin. Nature Cell Biol. 3, 107–113 (2001).

    Article  CAS  Google Scholar 

  17. Wohlschlegel, J.A. et al. Inhibition of eukaryotic DNA replication by geminin binding to Cdt1. Science 290, 2309–2312 (2000).

    Article  CAS  Google Scholar 

  18. Losada, A., Yokochi, T., Kobayashi, R. & Hirano, T. Identification and characterization of SA/Scc3p subunits in the Xenopus and human cohesin complexes. J. Cell Biol. 150, 405–416 (2000).

    Article  CAS  Google Scholar 

  19. Lohka, M.J. & Masui, Y. Formation in vitro of sperm pronuclei and mitotic chromosomes induced by amphibian ooplasmic components. Science 220, 719–721 (1983).

    Article  CAS  Google Scholar 

  20. Murray, A.W. Cell cycle extracts. Methods Cell Biol. 36, 581–605 (1991).

    Article  CAS  Google Scholar 

  21. Maiorano, D., Moreau, J. & Mechali, M. XCDT1 is required for the assembly of pre-replicative complexes in Xenopus laevis. Nature 404, 622–625 (2000).

    Article  CAS  Google Scholar 

  22. Krantz, I.D. et al. Cornelia de Lange syndrome is caused by mutations in NIPBL, the human homolog of Drosophila melanogaster Nipped-B. Nature Genet. 36, 631–635 (2004).

    Article  CAS  Google Scholar 

  23. Tonkin, E.T., Wang, T.J., Lisgo, S., Bamshad, M.J. & Strachan, T. NIPBL, encoding a homolog of fungal Scc2-type sister chromatid cohesion proteins and fly Nipped-B, is mutated in Cornelia de Lange syndrome. Nature Genet. 36, 636–641 (2004).

    Article  CAS  Google Scholar 

  24. Mendez, J. & Stillman, B. Chromatin association of human origin recognition complex, cdc6, and minichromosome maintenance proteins during the cell cycle: assembly of prereplication complexes in late mitosis. Mol. Cell. Biol. 20, 8602–8612 (2000).

    Article  CAS  Google Scholar 

  25. Walter, J. & Newport, J. Initiation of eukaryotic DNA replication: origin unwinding and sequential chromatin association of Cdc45, RPA, and DNA polymerase α. Mol. Cell 5, 617–627 (2000).

    Article  CAS  Google Scholar 

  26. Blow, J.J. Preventing re-replication of DNA in a single cell cycle: evidence for a replication licensing factor. J. Cell Biol. 122, 993–1002 (1993).

    Article  CAS  Google Scholar 

  27. Blow, J.J. & Laskey, R.A. Initiation of DNA replication in nuclei and purified DNA by a cell-free extract of Xenopus eggs. Cell 47, 577–587 (1986).

    Article  CAS  Google Scholar 

  28. O'Farrell, P.Z., Goodman, H.M. & O'Farrell, P.H. High resolution two-dimensional electrophoresis of basic as well as acidic proteins. Cell 12, 1133–1141 (1977).

    Article  CAS  Google Scholar 

  29. Peng, J. & Gygi, S.P. Proteomics: the move to mixtures. J. Mass Spectrom. 36, 1083–1091 (2001).

    Article  CAS  Google Scholar 

  30. Walter, J. & Newport, J.W. Regulation of replicon size in Xenopus egg extracts. Science 275, 993–995 (1997).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank T. Kelly, A. Dutta, D. Pellman, H. Masukata and the members of our laboratory for comments on the manuscript. We thank E. Arias for her gift of recombinant Cdt1 protein. This work was supported by National Institutes of Health grant GM62267 to J.W., and a Yamada Science Foundation fellowship (2002) and a Japan Society for the Promotion of Science postdoctoral fellowship (2004) to T.T.

Author information

Authors and Affiliations

  1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, 02115, MA, USA

    Tatsuro S. Takahashi, Pannyun Yiu & Johannes C. Walter

  2. Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, 02115, MA, USA

    Michael F. Chou & Steven Gygi

Authors
  1. Tatsuro S. Takahashi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Pannyun Yiu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Michael F. Chou
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Steven Gygi
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Johannes C. Walter
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Johannes C. Walter.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Cite this article

Takahashi, T., Yiu, P., Chou, M. et al. Recruitment of Xenopus Scc2 and cohesin to chromatin requires the pre-replication complex. Nat Cell Biol 6, 991–996 (2004). https://doi.org/10.1038/ncb1177

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ncb1177

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing