Letter | Published:

Selective Y centromere inactivation triggers chromosome shattering in micronuclei and repair by non-homologous end joining

Nature Cell Biology volume 19, pages 6875 (2017) | Download Citation

Abstract

Chromosome missegregation into a micronucleus can cause complex and localized genomic rearrangements1,2 known as chromothripsis3, but the underlying mechanisms remain unresolved. Here we developed an inducible Y centromere-selective inactivation strategy by exploiting a CENP-A/histone H3 chimaera to directly examine the fate of missegregated chromosomes in otherwise diploid human cells. Using this approach, we identified a temporal cascade of events that are initiated following centromere inactivation involving chromosome missegregation, fragmentation, and re-ligation that span three consecutive cell cycles. Following centromere inactivation, a micronucleus harbouring the Y chromosome is formed in the first cell cycle. Chromosome shattering, producing up to 53 dispersed fragments from a single chromosome, is triggered by premature micronuclear condensation prior to or during mitotic entry of the second cycle. Lastly, canonical non-homologous end joining (NHEJ), but not homology-dependent repair, is shown to facilitate re-ligation of chromosomal fragments in the third cycle. Thus, initial errors in cell division can provoke further genomic instability through fragmentation of micronuclear DNAs coupled to NHEJ-mediated reassembly in the subsequent interphase.

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Acknowledgements

We thank K. Jepsen and the UC San Diego IGM Genomics Center (MCC P30 CA023100) for DNA library preparation and sequencing, A. Shiau, S. Dowdy, E. Hatch, M. Hetzer, X. Wu and T. Pyntikova (Whitehead Institute, USA) for providing reagents, D. Jenkins, I. Goyal and Y. Sun for technical assistance, and the UC San Diego School of Medicine Microscopy Core (NINDS P30 NS047101) for shared use of equipment. This work was funded by a grant from the NIH (GM029513) to D.W.C., who receives salary support from the Ludwig Institute for Cancer Research. D.C.P. is supported by the Howard Hughes Medical Institute and NIH (HG007852). P.L. was supported by a Cancer Cell Biology Training Grant from the NCI (5T32CA067754-18) and a Postdoctoral Fellowship from the Hope Funds for Cancer Research (HFCR-14-06-06).

Author information

Author notes

    • Daniele Fachinetti

    Present address: Institut Curie, PSL Research University, CNRS, UMR144, F-75005, Paris, France.

Affiliations

  1. Ludwig Institute for Cancer Research and Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, California 92093, USA

    • Peter Ly
    • , Dong H. Kim
    • , Ofer Shoshani
    • , Daniele Fachinetti
    •  & Don W. Cleveland
  2. Department of Biology, Massachusetts Institute of Technology and Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA

    • Levi S. Teitz
    •  & David C. Page
  3. Howard Hughes Medical Institute and Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA

    • Helen Skaletsky
    •  & David C. Page

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Contributions

P.L. and D.W.C. conceived the project, designed the experiments, and wrote the manuscript. P.L. conducted the experiments and analysed the data. D.F. constructed the parental AID-tagged CENP-A cell line and provided key experimental input. P.L. and D.H.K. performed purification of micronuclei. O.S. assisted with FISH experiments. L.S.T., H.S. and D.C.P. analysed the sequencing data. All authors contributed comments on the final manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Don W. Cleveland.

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DOI

https://doi.org/10.1038/ncb3450

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