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Spontaneous occurrence of telomeric DNA damage response in the absence of chromosome fusions

Nature Structural & Molecular Biology volume 16, pages 1244–1251 (2009)Cite this article

Abstract

Telomere dysfunction is typically studied under conditions in which a component of the six-subunit shelterin complex that protects chromosome ends is disrupted. The nature of spontaneous telomere dysfunction is less well understood. Here we report that immortalized human cell lines lacking wild-type p53 function spontaneously show many telomeres with a DNA damage response (DDR), commonly affecting only one sister chromatid and not associated with increased chromosome end-joining. DDR+ telomeres represent an intermediate configuration between the fully capped and uncapped (fusogenic) states. In telomerase activity–positive (TA+) cells, DDR is associated with low TA and short telomeres. In cells using the alternative lengthening of telomeres mechanism (ALT+), DDR is partly independent of telomere length, mostly affects leading strand–replicated telomeres, and can be partly suppressed by TRF2 overexpression. In ALT+ (but not TA+) cells, DDR+ telomeres preferentially associate with large foci of extrachromosomal telomeric DNA and recombination proteins. DDR+ telomeres therefore arise through different mechanisms in TA+ and ALT+ cells and have different consequences.

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Figure 1: Spontaneous TIFs are present in untreated ALT+ cells.
Figure 2: Meta-TIFs are repressed in cell lines with wild-type p53 and high TA.
Figure 3: Distinct types of meta-TIFs are present in human cell lines.
Figure 4: Meta-TIFs in ALT+ cells result from telomere length–independent loss of function.
Figure 5: TRF2 suppresses telomere capping dysfunction in ALT+ cell lines.
Figure 6: Multiple dysfunctional telomeres in ALT+ cells colocalize with APB-like foci.
Figure 7: Three states of telomere end protection and a putative model to explain their occurrence.

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References

  1. Palm, W. & de Lange, T. How shelterin protects mammalian telomeres. Annu. Rev. Genet. 42, 301–334 (2008).

    Article  CAS  Google Scholar 

  2. Takai, H., Smogorzewska, A. & de Lange, T. DNA damage foci at dysfunctional telomeres. Curr. Biol. 13, 1549–1556 (2003).

    Article  CAS  Google Scholar 

  3. Denchi, E.L. & de Lange, T. Protection of telomeres through independent control of ATM and ATR by TRF2 and POT1. Nature 448, 1068–1071 (2007).

    Article  CAS  Google Scholar 

  4. Guo, X. et al. Dysfunctional telomeres activate an ATM-ATR-dependent DNA damage response to suppress tumorigenesis. EMBO J. 26, 4709–4719 (2007).

    Article  CAS  Google Scholar 

  5. Smogorzewska, A. & de Lange, T. Different telomere damage signaling pathways in human and mouse cells. EMBO J. 21, 4338–4348 (2002).

    Article  CAS  Google Scholar 

  6. Karlseder, J., Broccoli, D., Dai, Y., Hardy, S. & de Lange, T. p53- and ATM-dependent apoptosis induced by telomeres lacking TRF2. Science 283, 1321–1325 (1999).

    Article  CAS  Google Scholar 

  7. d'Adda di Fagagna, F. et al. A DNA damage checkpoint response in telomere-initiated senescence. Nature 426, 194–198 (2003).

    Article  CAS  Google Scholar 

  8. Smogorzewska, A., Karlseder, J., Holtgreve-Grez, H., Jauch, A. & de Lange, T. DNA ligase IV-dependent NHEJ of deprotected mammalian telomeres in G1 and G2. Curr. Biol. 12, 1635–1644 (2002).

    Article  CAS  Google Scholar 

  9. Colgin, L.M. & Reddel, R.R. Telomere maintenance mechanisms and cellular immortalization. Curr. Opin. Genet. Dev. 9, 97–103 (1999).

    Article  CAS  Google Scholar 

  10. Bryan, T.M., Englezou, A., Gupta, J., Bacchetti, S. & Reddel, R.R. Telomere elongation in immortal human cells without detectable telomerase activity. EMBO J. 14, 4240–4248 (1995).

    Article  CAS  Google Scholar 

  11. Bryan, T.M., Englezou, A., Dalla-Pozza, L., Dunham, M.A. & Reddel, R.R. Evidence for an alternative mechanism for maintaining telomere length in human tumors and tumor-derived cell lines. Nat. Med. 3, 1271–1274 (1997).

    Article  CAS  Google Scholar 

  12. Perrem, K. et al. Repression of an alternative mechanism for lengthening of telomeres in somatic cell hybrids. Oncogene 18, 3383–3390 (1999).

    Article  CAS  Google Scholar 

  13. Dunham, M.A., Neumann, A.A., Fasching, C.L. & Reddel, R.R. Telomere maintenance by recombination in human cells. Nat. Genet. 26, 447–450 (2000).

    Article  CAS  Google Scholar 

  14. Jiang, W.Q. et al. Suppression of alternative lengthening of telomeres by Sp100-mediated sequestration of MRE11/RAD50/NBS1 complex. Mol. Cell. Biol. 25, 2708–2721 (2005).

    Article  CAS  Google Scholar 

  15. Cesare, A.J. & Reddel, R.R. Telomere uncapping and alternative lengthening of telomeres. Mech. Ageing Dev. 129, 99–108 (2008).

    Article  CAS  Google Scholar 

  16. Wang, R.C., Smogorzewska, A. & de Lange, T. Homologous recombination generates T-loop-sized deletions at human telomeres. Cell 119, 355–368 (2004).

    Article  CAS  Google Scholar 

  17. Celli, G.B., Denchi, E.L. & de Lange, T. Ku70 stimulates fusion of dysfunctional telomeres yet protects chromosome ends from homologous recombination. Nat. Cell Biol. 8, 885–890 (2006).

    Article  Google Scholar 

  18. Wu, L. et al. Pot1 deficiency initiates DNA damage checkpoint activation and aberrant homologous recombination at telomeres. Cell 126, 49–62 (2006).

    Article  CAS  Google Scholar 

  19. Palm, W., Hockemeyer, D., Kibe, T. & de Lange, T. Functional dissection of human and mouse POT1 proteins. Mol. Cell. Biol. 29, 471–482 (2009).

    Article  CAS  Google Scholar 

  20. Cesare, A.J. & Griffith, J.D. Telomeric DNA in ALT cells is characterized by free telomeric circles and heterogeneous t-loops. Mol. Cell. Biol. 24, 9948–9957 (2004).

    Article  CAS  Google Scholar 

  21. Nabetani, A., Yokoyama, O. & Ishikawa, F. Localization of hRad9, hHus1, hRad1 and hRad17, and caffeine-sensitive DNA replication at ALT (alternative lengthening of telomeres)-associated promyelocytic leukemia body. J. Biol. Chem. 279, 25849–25857 (2004).

    Article  CAS  Google Scholar 

  22. Verdun, R.E., Crabbe, L., Haggblom, C. & Karlseder, J. Functional human telomeres are recognized as DNA damage in G2 of the cell cycle. Mol. Cell 20, 551–561 (2005).

    Article  CAS  Google Scholar 

  23. Nakamura, A.J. et al. Both telomeric and non-telomeric DNA damage are determinants of mammalian cellular senescence. Epigenetics Chromatin 1, 6 (2008).

    Article  Google Scholar 

  24. Anderson, M.J., Casey, G., Fasching, C.L. & Stanbridge, E.J. Evidence that wild-type TP53, and not genes on either chromosome I or II, controls the tumorigenic phenotype of the human fibrosarcoma HT1080. Genes Chromosom. Cancer 9, 266–281 (1994).

    Article  CAS  Google Scholar 

  25. Bunz, F. et al. Requirement for p53 and p21 to sustain G2 arrest after DNA damage. Science 282, 1497–1501 (1998).

    Article  CAS  Google Scholar 

  26. Cohen, S.B. & Reddel, R.R. A sensitive direct human telomerase activity assay. Nat. Methods 5, 355–360 (2008).

    Article  CAS  Google Scholar 

  27. Henson, J.D., Neumann, A.A., Yeager, T.R. & Reddel, R.R. Alternative lengthening of telomeres in mammalian cells. Oncogene 21, 598–610 (2002).

    Article  CAS  Google Scholar 

  28. Murnane, J.P., Sabatier, L., Marder, B.A. & Morgan, W.F. Telomere dynamics in an immortal human cell line. EMBO J. 13, 4953–4962 (1994).

    Article  CAS  Google Scholar 

  29. Bailey, S.M., Cornforth, M.N., Kurimasa, A., Chen, D.J. & Goodwin, E.H. Strand-specific postreplicative processing of mammalian telomeres. Science 293, 2462–2465 (2001).

    Article  CAS  Google Scholar 

  30. Yeager, T.R. et al. Telomerase-negative immortalized human cells contain a novel type of promyelocytic leukemia (PML) body. Cancer Res. 59, 4175–4179 (1999).

    CAS  PubMed  Google Scholar 

  31. Perrem, K., Colgin, L.M., Neumann, A.A., Yeager, T.R. & Reddel, R.R. Coexistence of alternative lengthening of telomeres and telomerase in hTERT-transfected GM847 cells. Mol. Cell. Biol. 21, 3862–3875 (2001).

    Article  CAS  Google Scholar 

  32. Pickett, H.A., Cesare, A.J., Johnstone, R.L., Neumann, A.A. & Reddel, R.R. Control of telomere length by a trimming mechanism that involves generation of t-circles. EMBO J. 28, 799–809 (2009).

    Article  CAS  Google Scholar 

  33. Sprung, C.N., Bryan, T.M., Reddel, R.R. & Murnane, J.P. Normal telomere maintenance in immortal ataxia telangiectasia cell lines. Mutat. Res. 379, 177–184 (1997).

    Article  CAS  Google Scholar 

  34. Bernardi, R. & Pandolfi, P.P. Structure, dynamics and functions of promyelocytic leukaemia nuclear bodies. Nat. Rev. Mol. Cell Biol. 8, 1006–1016 (2007).

    Article  CAS  Google Scholar 

  35. Griffith, J.D. et al. Mammalian telomeres end in a large duplex loop. Cell 97, 503–514 (1999).

    Article  CAS  Google Scholar 

  36. Stansel, R.M., de Lange, T. & Griffith, J.D. T-loop assembly in vitro involves binding of TRF2 near the 3′ telomeric overhang. EMBO J. 20, 5532–5540 (2001).

    Article  CAS  Google Scholar 

  37. Verdun, R.E. & Karlseder, J. Replication and protection of telomeres. Nature 447, 924–931 (2007).

    Article  CAS  Google Scholar 

  38. Bae, N.S. & Baumann, P.A. RAP1/TRF2 complex inhibits nonhomologous end-joining at human telomeric DNA ends. Mol. Cell 26, 323–334 (2007).

    Article  CAS  Google Scholar 

  39. Verdun, R.E. & Karlseder, J. The DNA damage machinery and homologous recombination pathway act consecutively to protect human telomeres. Cell 127, 709–720 (2006).

    Article  CAS  Google Scholar 

  40. Crabbe, L., Verdun, R.E., Haggblom, C.I. & Karlseder, J. Defective telomere lagging strand synthesis in cells lacking WRN helicase activity. Science 306, 1951–1953 (2004).

    Article  CAS  Google Scholar 

  41. Ding, H. et al. Regulation of murine telomere length by Rtel: an essential gene encoding a helicase-like protein. Cell 117, 873–886 (2004).

    Article  CAS  Google Scholar 

  42. Konishi, A. & de Lange, T. Cell cycle control of telomere protection and NHEJ revealed by a ts mutation in the DNA-binding domain of TRF2. Genes Dev. 22, 1221–1230 (2008).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank T. de Lange for supplying the TRF2ΔBΔM construct used in these experiments, and members of the Children's Medical Research Institute for comments on the manuscript. This work was supported by a US National Science Foundation international research fellowship (0602009), a grant from the Cure Cancer Australia Foundation and a Sir Keith Murdoch fellowship from the American Australian Association (to A.J.C.); a Promina postdoctoral fellowship (to H.A.P.); an Australian Postgraduate Award and a Judith Hyam Memorial Trust Fund for Cancer Research scholarship (to Z.K.) and a Cancer Council New South Wales program grant (to R.R.R.).

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Authors and Affiliations

  1. Cancer Research Unit, Children's Medical Research Institute, Westmead, New South Wales, Australia

    Anthony J Cesare, Zeenia Kaul, Scott B Cohen, Christine E Napier, Hilda A Pickett, Axel A Neumann & Roger R Reddel

  2. University of Sydney, Sydney, New South Wales, Australia

    Anthony J Cesare, Zeenia Kaul, Scott B Cohen, Christine E Napier, Hilda A Pickett, Axel A Neumann & Roger R Reddel

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  1. Anthony J Cesare
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  2. Zeenia Kaul
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Contributions

A.J.C., Z.K., S.B.C., C.E.N., H.A.P. and A.A.N. conducted the experiments. A.J.C, A.A.N. and R.R.R. designed the project. A.J.C. and R.R.R. analyzed the data and authored the manuscript.

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Correspondence to Roger R Reddel.

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Cesare, A., Kaul, Z., Cohen, S. et al. Spontaneous occurrence of telomeric DNA damage response in the absence of chromosome fusions. Nat Struct Mol Biol 16, 1244–1251 (2009). https://doi.org/10.1038/nsmb.1725

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