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.

Endonuclease-independent LINE-1 retrotransposition at mammalian telomeres

Abstract

Long interspersed element-1 (LINE-1 or L1) elements are abundant, non-long-terminal-repeat (non-LTR) retrotransposons that comprise 17% of human DNA1. The average human genome contains 80–100 retrotransposition-competent L1s (ref. 2), and they mobilize by a process that uses both the L1 endonuclease and reverse transcriptase, termed target-site primed reverse transcription3,4,5. We have previously reported an efficient, endonuclease-independent L1 retrotransposition pathway (ENi) in certain Chinese hamster ovary (CHO) cell lines that are defective in the non-homologous end-joining (NHEJ) pathway of DNA double-strand-break repair6. Here we have characterized ENi retrotransposition events generated in V3 CHO cells, which are deficient in DNA-dependent protein kinase catalytic subunit (DNA-PKcs) activity and have both dysfunctional telomeres and an NHEJ defect. Notably, 30% of ENi retrotransposition events insert in an orientation-specific manner adjacent to a perfect telomere repeat (5′-TTAGGG-3′). Similar insertions were not detected among ENi retrotransposition events generated in controls or in XR-1 CHO cells deficient for XRCC4, an NHEJ factor that is required for DNA ligation but has no known function in telomere maintenance. Furthermore, transient expression of a dominant-negative allele of human TRF2 (also called TERF2) in XRCC4-deficient XR-1 cells, which disrupts telomere capping, enables telomere-associated ENi retrotransposition events. These data indicate that L1s containing a disabled endonuclease can use dysfunctional telomeres as an integration substrate. The findings highlight similarities between the mechanism of ENi retrotransposition and the action of telomerase, because both processes can use a 3′ OH for priming reverse transcription at either internal DNA lesions or chromosome ends7,8. Thus, we propose that ENi retrotransposition is an ancestral mechanism of RNA-mediated DNA repair associated with non-LTR retrotransposons that may have been used before the acquisition of an endonuclease domain.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Characterization of L1 retrotransposition insertions in V3 cells.
Figure 2: EN i retrotransposition initiates at the telomere.
Figure 3: Destabilization of telomeres in XRCC4-deficient cells allows for telomere-associated EN i retrotransposition events.
Figure 4: Model for EN i retrotransposition in NHEJ-deficient cells containing dysfunctional telomeres.

References

  1. 1

    International Human Genome Sequencing Consortium. Initial sequencing and analysis of the human genome. Nature 409, 860–921 (2001)

  2. 2

    Brouha, B. et al. Hot L1s account for the bulk of retrotransposition in the human population. Proc. Natl Acad. Sci. USA 100, 5280–5285 (2003)

    ADS  CAS  Article  Google Scholar 

  3. 3

    Luan, D. D., Korman, M. H., Jakubczak, J. L. & Eickbush, T. H. Reverse transcription of R2Bm RNA is primed by a nick at the chromosomal target site: a mechanism for non-LTR retrotransposition. Cell 72, 595–605 (1993)

    CAS  Article  Google Scholar 

  4. 4

    Moran, J. V. et al. High frequency retrotransposition in cultured mammalian cells. Cell 87, 917–927 (1996)

    CAS  Article  Google Scholar 

  5. 5

    Feng, Q., Moran, J. V., Kazazian, H. H. & Boeke, J. D. Human L1 retrotransposon encodes a conserved endonuclease required for retrotransposition. Cell 87, 905–916 (1996)

    CAS  Article  Google Scholar 

  6. 6

    Morrish, T. A. et al. DNA repair mediated by endonuclease-independent LINE-1 retrotransposition. Nature Genet. 31, 159–165 (2002)

    CAS  Article  Google Scholar 

  7. 7

    Pardue, M. L. & DeBaryshe, P. G. Retrotransposons provide an evolutionarily robust non-telomerase mechanism to maintain telomeres. Annu. Rev. Genet. 37, 485–511 (2003)

    CAS  Article  Google Scholar 

  8. 8

    Lingner, J. et al. Reverse transcriptase motifs in the catalytic subunit of telomerase. Science 276, 561–567 (1997)

    CAS  Article  Google Scholar 

  9. 9

    Goytisolo, F. A., Samper, E., Edmonson, S., Taccioli, G. E. & Blasco, M. A. The absence of the DNA-dependent protein kinase catalytic subunit in mice results in anaphase bridges and in increased telomeric fusions with normal telomere length and G-strand overhang. Mol. Cell. Biol. 21, 3642–3651 (2001)

    CAS  Article  Google Scholar 

  10. 10

    Gilley, D. et al. DNA-PKcs is critical for telomere capping. Proc. Natl Acad. Sci. USA 98, 15084–15088 (2001)

    ADS  CAS  Article  Google Scholar 

  11. 11

    Bailey, S. M. et al. DNA double-strand break repair proteins are required to cap the ends of mammalian chromosomes. Proc. Natl Acad. Sci. USA 96, 14899–14904 (1999)

    ADS  CAS  Article  Google Scholar 

  12. 12

    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)

    ADS  CAS  Article  Google Scholar 

  13. 13

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

    CAS  Article  Google Scholar 

  14. 14

    Hao, L. Y., Strong, M. A. & Greider, C. W. Phosphorylation of H2AX at short telomeres in T cells and fibroblasts. J. Biol. Chem. 279, 45148–45154 (2004)

    CAS  Article  Google Scholar 

  15. 15

    de Lange, T. Shelterin: the protein complex that shapes and safeguards human telomeres. Genes Dev. 19, 2100–2110 (2005)

    CAS  Article  Google Scholar 

  16. 16

    Gilbert, N., Lutz-Prigge, S. & Moran, J. V. Genomic deletions created upon LINE-1 retrotransposition. Cell 110, 315–325 (2002)

    CAS  Article  Google Scholar 

  17. 17

    Gilbert, N., Lutz, S., Morrish, T. A. & Moran, J. V. Multiple fates of l1 retrotransposition intermediates in cultured human cells. Mol. Cell. Biol. 25, 7780–7795 (2005)

    CAS  Article  Google Scholar 

  18. 18

    Faravelli, M. et al. Molecular organization of internal telomeric sequences in Chinese hamster chromosomes. Gene 283, 11–16 (2002)

    CAS  Article  Google Scholar 

  19. 19

    Taccioli, G. E. et al. Targeted disruption of the catalytic subunit of the DNA-PK gene in mice confers severe combined immunodeficiency and radiosensitivity. Immunity 9, 355–366 (1998)

    CAS  Article  Google Scholar 

  20. 20

    Blunt, T. et al. Identification of a nonsense mutation in the carboxyl-terminal region of DNA-dependent protein kinase catalytic subunit in the scid mouse. Proc. Natl Acad. Sci. USA 93, 10285–10290 (1996)

    ADS  CAS  Article  Google Scholar 

  21. 21

    Whitmore, G. F., Varghese, A. J. & Gulyas, S. Cell cycle responses of two X-ray sensitive mutants defective in DNA repair. Int. J. Radiat. Biol. 56, 657–665 (1989)

    CAS  Article  Google Scholar 

  22. 22

    Blunt, T. et al. Defective DNA-dependent protein kinase activity is linked to V(D)J recombination and DNA repair defects associated with the murine scid mutation. Cell 80, 813–823 (1995)

    CAS  Article  Google Scholar 

  23. 23

    Meek, K., Gupta, S., Ramsden, D. A. & Lees-Miller, S. P. The DNA-dependent protein kinase: the director at the end. Immunol. Rev. 200, 132–141 (2004)

    CAS  Article  Google Scholar 

  24. 24

    Beamish, H. J. et al. The C-terminal conserved domain of DNA-PKcs, missing in the SCID mouse, is required for kinase activity. Nucleic Acids Res. 28, 1506–1513 (2000)

    CAS  Article  Google Scholar 

  25. 25

    van Steensel, B., Smogorzewska, A. & de Lange, T. TRF2 protects human telomeres from end-to-end fusions. Cell 92, 401–413 (1998)

    CAS  Article  Google Scholar 

  26. 26

    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)

    CAS  Article  Google Scholar 

  27. 27

    Celli, G. B. & de Lange, T. DNA processing is not required for ATM-mediated telomere damage response after TRF2 deletion. Nature Cell Biol. 7, 712–718 (2005)

    CAS  Article  Google Scholar 

  28. 28

    Kulpa, D. A. & Moran, J. V. Cis-preferential LINE-1 reverse transcriptase activity in ribonucleoprotein particles. Nature Struct. Mol. Biol. 13, 655–660 (2006)

    CAS  Article  Google Scholar 

  29. 29

    Wei, W., Morrish, T. A., Alisch, R. S. & Moran, J. V. A transient assay reveals that cultured human cells can accommodate multiple LINE-1 retrotransposition events. Anal. Biochem. 284, 435–438 (2000)

    CAS  Article  Google Scholar 

  30. 30

    Dombroski, B. A., Mathias, S. L., Nanthakumar, E., Scott, A. F. & Kazazian, H. H. Isolation of an active human transposable element. Science 254, 1805–1808 (1991)

    ADS  CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank A. M. DesLauriers for help with flow cytometry and cell sorting; R. Lyons for help with sequencing; T. de Lange for discussions and the dominant-negative TRF2 expression construct; M. Abe for the murine DNA-PKcs expression construct; G. Hammer and T. Else for help with the metaphase analysis; and C. Greider for discussions and use of equipment and reagents for immunocytochemistry experiments. We thank T. Else, T. Glover, T. Glaser, N. Howlett, T. Wilson and current members of the Moran laboratory for discussions during the course of this work. T.A.M. was supported in part by a NIH training grant and is now a Leukemia and Lymphoma Society Fellow. J.L.G.-P. was supported in part by a MEC/Fulbright postdoctoral grant. G.E.T. is partially supported by a grant from the Human Frontier Science Program. T.D.S was supported in part by a grant from the National Institutes of Health. J.S. was supported in part by a grant from the Pew Foundation. J.V.M was supported in part by grants from the W. M. Keck Foundation and the National Institutes of Health.

Author Contributions T.A.M is co-first author and is a corresponding author. She contributed to the original concept, designed and performed experiments, analysed the data, and wrote the first draft of the manuscript. J.L.G.-P. is co-first author. He contributed to the concept, designed and performed experiments, analysed the data, and helped write and revise the manuscript. T.D.S and G.E.T. contributed reagents, helped with data interpretation and provided helpful comments during the course of this study. J.S. contributed the murine DNA-PKcs complemented cell lines, performed functional analyses on those cell lines, helped with data analysis, and helped revise the manuscript. J.V.M. is the senior author. He contributed to the original concept, analysed the data, revised the manuscript and provided financial support.

Author information

Affiliations

Authors

Corresponding authors

Correspondence to Tammy A. Morrish or John V. Moran.

Ethics declarations

Competing interests

Reprints and permissions information is available at www.nature.com/reprints. The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-4 with Legends, Supplementary Methods, Supplementary Tables 1-3 and additional references (PDF 1675 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Morrish, T., Garcia-Perez, J., Stamato, T. et al. Endonuclease-independent LINE-1 retrotransposition at mammalian telomeres. Nature 446, 208–212 (2007). https://doi.org/10.1038/nature05560

Download citation

Further reading

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