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.

  • Progress
  • Published:

Pathways of mammalian replication fork restart

Abstract

Single-molecule analyses of DNA replication have greatly advanced our understanding of mammalian replication restart. Several proteins that are not part of the core replication machinery promote the efficient restart of replication forks that have been stalled by replication inhibitors, suggesting that bona fide fork restart pathways exist in mammalian cells. Different models of replication fork restart can be envisaged, based on the involvement of DNA helicases, nucleases, homologous recombination factors and the importance of DNA double-strand break formation.

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

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

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

Figure 1: Models of replication fork restart.
Figure 2: Early replication fork restart and late replication fork repair.

Similar content being viewed by others

References

  1. Hanada, K. et al. The structure-specific endonuclease Mus81 contributes to replication restart by generating double-strand DNA breaks. Nature Struct. Mol. Biol. 14, 1096–1104 (2007).

    Article  CAS  Google Scholar 

  2. Heller, R. C. & Marians, K. J. Replisome assembly and the direct restart of stalled replication forks. Nature Rev. Mol. Cell Biol. 7, 932–943 (2006).

    Article  CAS  Google Scholar 

  3. Ibarra, A., Schwob, E. & Mendez, J. Excess MCM proteins protect human cells from replicative stress by licensing backup origins of replication. Proc. Natl Acad. Sci. USA 105, 8956–8961 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Ge, X. Q., Jackson, D. A. & Blow, J. J. Dormant origins licensed by excess MCM2–7 are required for human cells to survive replicative stress. Genes Dev. 21, 3331–3341 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Lundin, C. et al. Different roles for nonhomologous end joining and homologous recombination following replication arrest in mammalian cells. Mol. Cell. Biol. 22, 5869–5878 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Saintigny, Y. et al. Characterization of homologous recombination induced by replication inhibition in mammalian cells. Embo J. 20, 3861–3870 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Tuduri, S., Tourriere, H. & Pasero, P. Defining replication origin efficiency using DNA fiber assays. Chromosome Res. 18, 91–102 (2010).

    Article  CAS  PubMed  Google Scholar 

  8. Davies, S. L., North, P. S. & Hickson, I. D. Role for BLM in replication-fork restart and suppression of origin firing after replicative stress. Nature Struct. Mol. Biol. 14, 677–679 (2007).

    Article  CAS  Google Scholar 

  9. Franchitto, A. et al. Replication fork stalling in WRN-deficient cells is overcome by prompt activation of a MUS81-dependent pathway. J. Cell Biol. 183, 241–252 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Sidorova, J. M., Li, N., Folch, A. & Monnat, R. J. Jr. The RecQ helicase WRN is required for normal replication fork progression after DNA damage or replication fork arrest. Cell Cycle 7, 796–807 (2008).

    Article  CAS  PubMed  Google Scholar 

  11. Bryant, H. E. et al. PARP is activated at stalled forks to mediate Mre11-dependent replication restart and recombination. Embo J. 28, 2601–2615 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Ciccia, A. et al. The SIOD disorder protein SMARCAL1 is an RPA-interacting protein involved in replication fork restart. Genes Dev. 23, 2415–2425 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Petermann, E., Orta, M. L., Issaeva, N., Schultz, N. & Helleday, T. Hydroxyurea-stalled replication forks become progressively inactivated and require two different RAD51-mediated pathways for restart and repair. Mol. Cell 37, 492–502 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Liu, J., Xu, L., Sandler, S. J. & Marians, K. J. Replication fork assembly at recombination intermediates is required for bacterial growth. Proc. Natl Acad. Sci. USA 96, 3552–3555 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Llorente, B., Smith, C. E. & Symington, L. S. Break-induced replication: what is it and what is it for? Cell Cycle 7, 859–864 (2008).

    Article  CAS  PubMed  Google Scholar 

  16. Karow, J. K., Constantinou, A., Li, J. L., West, S. C. & Hickson, I. D. The Bloom's syndrome gene product promotes branch migration of holliday junctions. Proc. Natl Acad. Sci. USA 97, 6504–6508 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Ralf, C., Hickson, I. D. & Wu, L. The Bloom's syndrome helicase can promote the regression of a model replication fork. J. Biol. Chem. 281, 22839–22846 (2006).

    Article  CAS  PubMed  Google Scholar 

  18. Bugreev, D. V., Yu, X., Egelman, E. H. & Mazin, A. V. Novel pro- and anti-recombination activities of the Bloom's syndrome helicase. Genes Dev. 21, 3085–3094 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Rassool, F. V., North, P. S., Mufti, G. J. & Hickson, I. D. Constitutive DNA damage is linked to DNA replication abnormalities in Bloom's syndrome cells. Oncogene 22, 8749–8757 (2003).

    Article  CAS  PubMed  Google Scholar 

  20. Constantinou, A. et al. Werner's syndrome protein (WRN) migrates Holliday junctions and co-localizes with RPA upon replication arrest. EMBO Rep. 1, 80–84 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Machwe, A., Xiao, L., Groden, J. & Orren, D. K. The Werner and Bloom syndrome proteins catalyze regression of a model replication fork. Biochemistry 45, 13939–13946 (2006).

    Article  CAS  PubMed  Google Scholar 

  22. Sidorova, J. M. Roles of the Werner syndrome RecQ helicase in DNA replication. DNA Repair (Amst.) 7, 1776–1786 (2008).

    Article  CAS  Google Scholar 

  23. Yusufzai, T. & Kadonaga, J. T. HARP is an ATP-driven annealing helicase. Science 322, 748–750 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Bansbach, C. E., Betous, R., Lovejoy, C. A., Glick, G. G. & Cortez, D. The annealing helicase SMARCAL1 maintains genome integrity at stalled replication forks. Genes Dev. 23, 2405–2414 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Baumann, P., Benson, F. E. & West, S. C. Human Rad51 protein promotes ATP-dependent homologous pairing and strand transfer reactions in vitro. Cell 87, 757–766 (1996).

    Article  CAS  PubMed  Google Scholar 

  26. Seigneur, M., Ehrlich, S. D. & Michel, B. RuvABC-dependent double-strand breaks in DnaBts mutants require RecA. Mol. Microbiol. 38, 565–574 (2000).

    Article  CAS  PubMed  Google Scholar 

  27. Robu, M. E., Inman, R. B. & Cox, M. M. RecA protein promotes the regression of stalled replication forks in vitro. Proc. Natl Acad. Sci. USA 98, 8211–8218 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Yoon, D., Wang, Y., Stapleford, K., Wiesmuller, L. & Chen, J. p53 inhibits strand exchange and replication fork regression promoted by human Rad51. J. Mol. Biol. 336, 639–654 (2004).

    Article  CAS  PubMed  Google Scholar 

  29. Buis, J. et al. Mre11 nuclease activity has essential roles in DNA repair and genomic stability distinct from ATM activation. Cell 135, 85–96 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Sartori, A. A. et al. Human CtIP promotes DNA end resection. Nature 450, 509–514 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Franchitto, A. & Pichierri, P. Bloom's syndrome protein is required for correct relocalization of RAD50/MRE11/NBS1 complex after replication fork arrest. J. Cell Biol. 157, 19–30 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Franchitto, A. & Pichierri, P. Werner syndrome protein and the MRE11 complex are involved in a common pathway of replication fork recovery. Cell Cycle 3, 1331–1339 (2004).

    Article  CAS  PubMed  Google Scholar 

  33. Tittel-Elmer, M., Alabert, C., Pasero, P. & Cobb, J. A. The MRX complex stabilizes the replisome independently of the S phase checkpoint during replication stress. Embo J. 28, 1142–1156 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Terret, M. E., Sherwood, R., Rahman, S., Qin, J. & Jallepalli, P. V. Cohesin acetylation speeds the replication fork. Nature 462, 231–234 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Bishop, D. K. et al. Xrcc3 is required for assembly of Rad51 complexes in vivo. J. Biol. Chem. 273, 21482–21488 (1998).

    Article  CAS  PubMed  Google Scholar 

  36. Raynard, S., Bussen, W. & Sung, P. A double Holliday junction dissolvasome comprising BLM, topoisomerase IIIα, and BLAP75. J. Biol. Chem. 281, 13861–13864 (2006).

    Article  CAS  PubMed  Google Scholar 

  37. Singh, T. R. et al. BLAP18/RMI2, a novel OB-fold-containing protein, is an essential component of the Bloom helicase-double Holliday junction dissolvasome. Genes Dev. 22, 2856–2868 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Wu, L. & Hickson, I. D. The Bloom's syndrome helicase suppresses crossing over during homologous recombination. Nature 426, 870–874 (2003).

    Article  CAS  PubMed  Google Scholar 

  39. Shimura, T. et al. Bloom's syndrome helicase and Mus81 are required to induce transient double-strand DNA breaks in response to DNA replication stress. J. Mol. Biol. 375, 1152–1164 (2008).

    Article  CAS  PubMed  Google Scholar 

  40. Malkova, A., Naylor, M. L., Yamaguchi, M., Ira, G. & Haber, J. E. RAD51-dependent break-induced replication differs in kinetics and checkpoint responses from RAD51-mediated gene conversion. Mol. Cell Biol. 25, 933–944 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Lee, S. H. et al. The SET domain protein Metnase mediates foreign DNA integration and links integration to nonhomologous end-joining repair. Proc. Natl Acad. Sci. USA 102, 18075–18080 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. De Haro, L. P. et al. Metnase promotes restart and repair of stalled and collapsed replication forks. Nucleic Acids Res. 10 May 2010 (doi:10.1093/nar/gkq339).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Feijoo, C. et al. Activation of mammalian Chk1 during DNA replication arrest: a role for Chk1 in the intra-S phase checkpoint monitoring replication origin firing. J. Cell Biol. 154, 913–923 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Zachos, G., Rainey, M. D. & Gillespie, D. A. Chk1-dependent S-M checkpoint delay in vertebrate cells is linked to maintenance of viable replication structures. Mol. Cell Biol. 25, 563–574 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Scorah, J. & McGowan, C. H. Claspin and Chk1 regulate replication fork stability by different mechanisms. Cell Cycle 8, 1036–1043 (2009).

    Article  CAS  PubMed  Google Scholar 

  46. Leman, A. R., Noguchi, C., Lee, C. Y. & Noguchi, E. Human Timeless and Tipin stabilize replication forks and facilitate sister-chromatid cohesion. J. Cell Sci. 123, 660–670 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Tanaka, H. et al. Replisome progression complex links DNA replication to sister chromatid cohesion in Xenopus egg extracts. Genes Cells 14, 949–963 (2009).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank the Medical Research Council, the Swedish Research Council, the Swedish Children's Cancer Foundation, the Swedish Pain Relief Foundation and the Swedish Cancer Society for financial support.

Author information

Authors and Affiliations

Authors

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary information 1 (box)

Replication fork restart in E. coli (PDF 76 kb)

Supplementary information 2 (box)

Chromosome combing and DNA fiber technique (PDF 59 kb)

Supplementary information 3 (box)

Replication fork stabilisation (PDF 80 kb)

Related links

Related links

FURTHER INFORMATION

Thomas Helleday's homepage

Glossary

Aphidicolin

A small-molecule inhibitor that directly blocks the activity of the replicative DNA polymerases. Aphidicolin treatment leads to replication fork stalling and eventually DNA DSB formation.

Break-induced replication

A mechanism for origin-independent replication restart, whereby a resected DNA end invades a homologous DNA molecule, thus establishing a replication fork.

Chromosome combing

A method for single-molecule analysis of DNA replication forks. Newly replicated DNA is labelled in vivo using halogenated thymidine analogues and genomic DNA is isolated and spread out on microscope coverslips. Immunofluorescence staining of the thymidine analogues is used to visualize the labelled tracks that are left on the DNA by moving replication forks.

Core replication machinery

The complex of proteins that is essential for all DNA replication and includes the replicative DNA helicase, primase, clamp loader, sliding clamp and leading- and lagging-strand DNA polymerases.

DNA fibre technique

A technique that is similar to chromosome combing, but in which cells on microscope slides are treated with detergent and the DNA is spread directly out of the lysed nuclei. Because of the lysis method, this technique is used in vertebrate cells but not in yeast.

DNA helicase

An enzyme that translocates on DNA and unwinds the double helix into ssDNA in an ATP-driven reaction. Annealing helicases use ATP to catalyse the reverse reaction.

Holliday junction

A four-way junction between two dsDNA molecules of homologous sequence. Holliday junctions are mobile and can be translocated by DNA helicases (branch migration).

Hydroxyurea

A radical scavenger that inhibits ribonucleotide reductase, which results in cells producing less of the desoxyribonucleotides that are used for DNA synthesis. Hydroxyurea treatment leads to replication fork stalling and eventually DNA DSB formation.

Lagging strand

The nascent strand of DNA that is synthesized discontinuously in short pieces (Okazaki fragments) at the replication fork.

Origin firing

The start of replication fork progression at a replication origin. Mammalian origin firing is restricted to S phase and is controlled by cell cycle signalling.

Replication fork

A structure formed when the template strands have been separated by helicases and a newly formed copy of the DNA is synthesized. The fork moves in the direction of leading-strand synthesis.

Replication fork restart

The resumption of fork progression after removal or bypass of a replication block.

Replication fork stabilization

The maintenance of viable replication fork structures and the replication machinery during a replication block.

Replication origin

The chromosomal location at which new replication factories are assembled and replication is initiated. Replication fork movement from origins is bidirectional.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Petermann, E., Helleday, T. Pathways of mammalian replication fork restart. Nat Rev Mol Cell Biol 11, 683–687 (2010). https://doi.org/10.1038/nrm2974

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nrm2974

This article is cited by

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