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.

Cell-type-specific replication initiation programs set fragility of the FRA3B fragile site

Abstract

Common fragile sites have long been identified by cytogeneticists as chromosomal regions prone to breakage upon replication stress1. They are increasingly recognized to be preferential targets for oncogene-induced DNA damage in pre-neoplastic lesions2 and hotspots for chromosomal rearrangements in various cancers3. Common fragile site instability was attributed to the fact that they contain sequences prone to form secondary structures that may impair replication fork movement, possibly leading to fork collapse resulting in DNA breaks4. Here we show, in contrast to this view, that the fragility of FRA3B—the most active common fragile site in human lymphocytes—does not rely on fork slowing or stalling but on a paucity of initiation events. Indeed, in lymphoblastoid cells, but not in fibroblasts, initiation events are excluded from a FRA3B core extending approximately 700 kilobases, which forces forks coming from flanking regions to cover long distances in order to complete replication. We also show that origins of the flanking regions fire in mid-S phase, leaving the site incompletely replicated upon fork slowing. Notably, FRA3B instability is specific to cells showing this particular initiation pattern. The fact that both origin setting5,6 and replication timing are highly plastic7,8 in mammalian cells explains the tissue specificity of common fragile site instability we observed. Thus, we propose that common fragile sites correspond to the latest initiation-poor regions to complete replication in a given cell type. For historical reasons, common fragile sites have been essentially mapped in lymphocytes1. Therefore, common fragile site contribution to chromosomal rearrangements in tumours should be reassessed after mapping fragile sites in the cell type from which each tumour originates.

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: Comparison of fork properties in the FHIT locus and in the bulk genome in JEFF cells.
Figure 2: Mapping of initiation and termination events along the FHIT locus.
Figure 3: Relationship between replication profile of the FHIT locus and FRA3B fragility in lymphoblastoid and fibroblastic cells.

References

  1. Sutherland, G. R. & Richards, R. I. The molecular basis of fragile sites in human chromosomes. Curr. Opin. Genet. Dev. 5, 323–327 (1995)

    Article  CAS  Google Scholar 

  2. Negrini, S., Gorgoulis, V. G. & Halazonetis, T. D. Genomic instability—an evolving hallmark of cancer. Nature Rev. Mol. Cell Biol. 11, 220–228 (2010)

    Article  CAS  Google Scholar 

  3. Bignell, G. R. et al. Signatures of mutation and selection in the cancer genome. Nature 463, 893–898 (2010)

    Article  ADS  CAS  Google Scholar 

  4. Schwartz, M., Zlotorynski, E. & Kerem, B. The molecular basis of common and rare fragile sites. Cancer Lett. 232, 13–26 (2006)

    Article  CAS  Google Scholar 

  5. Grégoire, D., Brodolin, K. & Méchali, M. HoxB domain induction silences DNA replication origins in the locus and specifies a single origin at its boundary. EMBO Rep. 7, 812–816 (2006)

    PubMed  PubMed Central  Google Scholar 

  6. Dazy, S., Gandrillon, O., Hyrien, O. & Prioleau, M. N. Broadening of DNA replication origin usage during metazoan cell differentiation. EMBO Rep. 7, 806–811 (2006)

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Hansen, R. S. et al. Sequencing newly replicated DNA reveals widespread plasticity in human replication timing. Proc. Natl Acad. Sci. USA 107, 139–144 (2010)

    Article  ADS  CAS  Google Scholar 

  8. Hiratani, I. et al. Global reorganization of replication domains during embryonic stem cell differentiation. PLoS Biol. 6, e245 (2008)

    Article  Google Scholar 

  9. Tourriere, H. & Pasero, P. Maintenance of fork integrity at damaged DNA and natural pause sites. DNA Repair 6, 900–913 (2007)

    Article  CAS  Google Scholar 

  10. Branzei, D. & Foiani, M. Maintaining genome stability at the replication fork. Nature Rev. Mol. Cell Biol. 11, 208–219 (2010)

    Article  CAS  Google Scholar 

  11. Durkin, S. G. & Glover, T. W. Chromosome fragile sites. Annu. Rev. Genet. 41, 169–192 (2007)

    Article  CAS  Google Scholar 

  12. Helmrich, A., Stout-Weider, K., Hermann, K., Schrock, E. & Heiden, T. Common fragile sites are conserved features of human and mouse chromosomes and relate to large active genes. Genome Res. 16, 1222–1230 (2006)

    Article  CAS  Google Scholar 

  13. Tsantoulis, P. K. et al. Oncogene-induced replication stress preferentially targets common fragile sites in preneoplastic lesions. A genome-wide study. Oncogene 27, 3256–3264 (2008)

    Article  CAS  Google Scholar 

  14. Palumbo, E., Matricardi, L., Tosoni, E., Bensimon, A. & Russo, A. Replication dynamics at common fragile site FRA6E . Chromosoma (2010)

  15. Huebner, K. & Croce, C. M. Cancer and the FRA3B/FHIT fragile locus: it’s a HIT. Br. J. Cancer 88, 1501–1506 (2003)

    Article  CAS  Google Scholar 

  16. Pichiorri, F. et al. Molecular parameters of genome instability: roles of fragile genes at common fragile sites. J. Cell. Biochem. 104, 1525–1533 (2008)

    Article  CAS  Google Scholar 

  17. Lebofsky, R., Heilig, R., Sonnleitner, M., Weissenbach, J. & Bensimon, A. DNA replication origin interference increases the spacing between initiation events in human cells. Mol. Biol. Cell 17, 5337–5345 (2006)

    Article  CAS  Google Scholar 

  18. Cha, R. S. & Kleckner, N. ATR homolog Mec1 promotes fork progression, thus averting breaks in replication slow zones. Science 297, 602–606 (2002)

    Article  ADS  CAS  Google Scholar 

  19. Rothstein, R., Michel, B. & Gangloff, S. Replication fork pausing and recombination or “gimme a break”. Genes Dev. 14, 1–10 (2000)

    CAS  PubMed  Google Scholar 

  20. Farkash-Amar, S. et al. Global organization of replication time zones of the mouse genome. Genome Res. 18, 1562–1570 (2008)

    Article  CAS  Google Scholar 

  21. Anglana, M., Apiou, F., Bensimon, A. & Debatisse, M. Dynamics of DNA replication in mammalian somatic cells: nucleotide pool modulates origin choice and interorigin spacing. Cell 114, 385–394 (2003)

    Article  CAS  Google Scholar 

  22. Courbet, S. et al. Replication fork movement sets chromatin loop size and origin choice in mammalian cells. Nature 455, 557–560 (2008)

    Article  ADS  CAS  Google Scholar 

  23. Bielinsky, A. K. Replication origins: why do we need so many? Cell Cycle 2, 307–309 (2003)

    Article  CAS  Google Scholar 

  24. El Achkar, E., Gerbault-Seureau, M., Muleris, M., Dutrillaux, B. & Debatisse, M. Premature condensation induces breaks at the interface of early and late replicating chromosome bands bearing common fragile sites. Proc. Natl Acad. Sci. USA 102, 18069–18074 (2005)

    Article  ADS  CAS  Google Scholar 

  25. Le Beau, M. M. et al. Replication of a common fragile site, FRA3B, occurs late in S phase and is delayed further upon induction: implications for the mechanism of fragile site induction. Hum. Mol. Genet. 7, 755–761 (1998)

    Article  CAS  Google Scholar 

  26. Durkin, S. G. et al. Replication stress induces tumor-like microdeletions in FHIT/FRA3B. Proc. Natl Acad. Sci. USA 105, 246–251 (2008)

    Article  ADS  CAS  Google Scholar 

  27. Ryba, T. et al. Evolutionarily conserved replication timing profiles predict long-range chromatin interactions and distinguish closely related cell types. Genome Res. 20, 761–770 (2010)

    Article  CAS  Google Scholar 

  28. O’Keefe, L. V. & Richards, R. I. Common chromosomal fragile sites and cancer: focus on FRA16D. Cancer Lett. 232, 37–47 (2006)

    Article  Google Scholar 

  29. Masai, H., Matsumoto, S., You, Z., Yoshizawa-Sugata, N. & Oda, M. Eukaryotic chromosome DNA replication: where, when, and how? Annu. Rev. Biochem. 79, 89–130 (2010)

    Article  CAS  Google Scholar 

  30. Michalet, X. et al. Dynamic molecular combing: stretching the whole human genome for high-resolution studies. Science 277, 1518–1523 (1997)

    Article  CAS  Google Scholar 

  31. Labit, H. et al. A simple and optimized method of producing silanized surfaces for FISH and replication mapping on combed DNA fibers. Biotechniques 45, 649–658 (2008)

    Article  CAS  Google Scholar 

  32. R Development Core Team. R: A Language and Environment for Statistical Computing (R Foundation for Statistical Computing, 2006)

Download references

Acknowledgements

We thank E. Blackburn for critical reading of the manuscript. We thank Genomic Vision for making the DNA combing technology available to us. We acknowledge the Nikon Imaging Centre at Institut Curie-CNRS. We thank C. Rouzaud for help in combing experiments. A.L. is supported by a grant from the ARC (Association pour la recherche sur le cancer). The M.D. team is supported by La Ligue Nationale contre le Cancer (LNCC) (Equipe Labellisée EL2008.LNCC/MD), INCa (Institut National du Cancer) (2009-1-PLBIO-10-IC-1) and the PIC Réplication, Instabilité Chromosomique et Cancer (Institut Curie).

Author information

Authors and Affiliations

Authors

Contributions

A.L. performed and analysed combing experiments. G.A.M. performed statistical and Repli-Seq analyses. S.K., A.-M.L. and O.B. performed cytogenetic analyses. N.V. and B.M. designed the Morse code. R.S.H. contributed to Repli-Seq analysis. A.L., G.A.M., O.B. and M.D. wrote the paper. M.D. planned the project.

Corresponding author

Correspondence to Michelle Debatisse.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

The file contains Supplementary Figures 1-9 with legends and Supplementary Tables 1-4. (PDF 2012 kb)

PowerPoint slides

Rights and permissions

Reprints and permissions

About this article

Cite this article

Letessier, A., Millot, G., Koundrioukoff, S. et al. Cell-type-specific replication initiation programs set fragility of the FRA3B fragile site. Nature 470, 120–123 (2011). https://doi.org/10.1038/nature09745

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature09745

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

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