Abstract

Defined transcription factors can induce epigenetic reprogramming of adult mammalian cells into induced pluripotent stem cells. Although DNA factors are integrated during some reprogramming methods, it is unknown whether the genome remains unchanged at the single nucleotide level. Here we show that 22 human induced pluripotent stem (hiPS) cell lines reprogrammed using five different methods each contained an average of five protein-coding point mutations in the regions sampled (an estimated six protein-coding point mutations per exome). The majority of these mutations were non-synonymous, nonsense or splice variants, and were enriched in genes mutated or having causative effects in cancers. At least half of these reprogramming-associated mutations pre-existed in fibroblast progenitors at low frequencies, whereas the rest occurred during or after reprogramming. Thus, hiPS cells acquire genetic modifications in addition to epigenetic modifications. Extensive genetic screening should become a standard procedure to ensure hiPS cell safety before clinical use.

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Data deposits

Sequencing results for the mutations reported here are included in Supplementary Figure 1. Raw Illumina sequencing reads are available from the NCBI ShortRead Archive, accession SRP005709, except for lines derived from Hib11, Hib17, Hib29, CF, HFFxF, dH1F fibroblasts as the original donors were not consulted about public release of their genome data.

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Acknowledgements

We thank J. M. Akey, G. M. Church, S. Ding, J. B. Li and J. Shendure for discussions and suggestions, S. Vassallo for assistance with DNA shearing, and G. L. Boulting and S. Ratansirintrawoot for assistance with hiPS cell culture. This study is supported by NIH R01 HL094963 and a UCSD new faculty start-up fund (to K.Z.), a training grant from the California Institute for Regenerative Medicine (TG2-01154) and a CIRM grant (RC1-00116) (to L.S.B.G.). L.S.B.G. is an Investigator of the Howard Hughes Medical Institute. A. Gore is supported by the Focht-Powell Fellowship and a CIRM predoctoral fellowship. M.L.W. is supported by an institutional training grant from the National Institute of General Medical Sciences (T32 GM008666). Y.-H.L. is supported by the A*Star Institute of Medical Biology and the Singapore Stem Cell Consortium. Work in the laboratory of J.C.I.B. was supported by grants from MICINN, Sanofi-Aventis, the G. Harold and Leila Y. Mathers Foundation and the Cellex Foundation. G.Q.D. is an investigator of the Howard Hughes Medical Institute and supported by grants from the NIH.

Author information

Author notes

    • Athurva Gore
    •  & Zhe Li

    These authors contributed equally to this work.

Affiliations

  1. Department of Bioengineering, Institute for Genomic Medicine and Institute of Engineering in Medicine, University of California at San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA

    • Athurva Gore
    • , Zhe Li
    • , Ho-Lim Fung
    •  & Kun Zhang
  2. Department of Cellular and Molecular Medicine and Howard Hughes Medical Institute, University of California at San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA

    • Jessica E. Young
    • , Isabel Canto
    • , Mason A. Israel
    • , Melissa L. Wilbert
    •  & Lawrence S. B. Goldstein
  3. Division of Pediatric Hematology/Oncology, Children’s Hospital Boston and Dana Farber Cancer Institute, Boston, Massachusetts 02115, USA

    • Suneet Agarwal
    • , Yuin-Han Loh
    • , Philip D. Manos
    •  & George Q. Daley
  4. Department of Anatomy, University of Wisconsin-Madison, Madison, Wisconsin 53705, USA

    • Jessica Antosiewicz-Bourget
    • , Junying Yu
    •  & James A. Thomson
  5. Center of Regenerative Medicine, 08003 Barcelona, Spain

    • Alessandra Giorgetti
    • , Nuria Montserrat
    •  & Juan Carlos Izpisua Belmonte
  6. Howard Hughes Medical Institute, Harvard Stem Cell Institute, Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, Massachusetts 02138, USA

    • Evangelos Kiskinis
    •  & Kevin Eggan
  7. Department of Genetics, Harvard Medical School, Boston, Massachusetts 02135, USA

    • Je-Hyuk Lee
  8. Salk Institute for Biological Studies, La Jolla, California 92037, USA

    • Athanasia D. Panopoulos
    • , Sergio Ruiz
    •  & Juan Carlos Izpisua Belmonte
  9. The J. Craig Venter Institute, Rockville, Maryland 20850, USA

    • Ewen F. Kirkness
  10. Immune Disease Institute, Children’s Hospital Boston, Boston, Massachusetts 02115, USA

    • Derrick J. Rossi

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Contributions

L.S.B.G. and K.Z. co-directed the study. A. Gore, Z.L., L.S.B.G. and K.Z. designed the experiments. J.E.Y., S.A., J.A.-B., I.C., A. Giorgetti, M.A.I., E.K., J.-H.L., Y.-H.L., P.D.M., N.M., A.D.P., S.R., M.L.W., J. Yu, J.C.I.B., D.J.R., J.A.T., K.E., G.Q.D. and L.S.B.G. biopsied, cultured and derived hiPS cell lines. Z.L. performed DNA extraction. A. Gore, Z.L. and K.Z. performed exome library construction, DigiQ library construction and validation Sanger sequencing. H.-L.F. performed Illumina sequencing. A. Gore and K.Z. performed bioinformatic and statistical analysis with contributions from E.F.K. A. Gore, Z.L., L.S.B.G., G.Q.D. and K.Z. wrote the manuscript with contributions from all other authors.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Lawrence S. B. Goldstein or Kun Zhang.

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https://doi.org/10.1038/nature09805

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