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Mitochondrial gene replacement in primate offspring and embryonic stem cells

Nature volume 461, pages 367372 (17 September 2009) | Download Citation

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  • A Corrigendum to this article was published on 10 December 2014

Abstract

Mitochondria are found in all eukaryotic cells and contain their own genome (mitochondrial DNA or mtDNA). Unlike the nuclear genome, which is derived from both the egg and sperm at fertilization, the mtDNA in the embryo is derived almost exclusively from the egg; that is, it is of maternal origin. Mutations in mtDNA contribute to a diverse range of currently incurable human diseases and disorders. To establish preclinical models for new therapeutic approaches, we demonstrate here that the mitochondrial genome can be efficiently replaced in mature non-human primate oocytes (Macaca mulatta) by spindle–chromosomal complex transfer from one egg to an enucleated, mitochondrial-replete egg. The reconstructed oocytes with the mitochondrial replacement were capable of supporting normal fertilization, embryo development and produced healthy offspring. Genetic analysis confirmed that nuclear DNA in the three infants born so far originated from the spindle donors whereas mtDNA came from the cytoplast donors. No contribution of spindle donor mtDNA was detected in offspring. Spindle replacement is shown here as an efficient protocol replacing the full complement of mitochondria in newly generated embryonic stem cell lines. This approach may offer a reproductive option to prevent mtDNA disease transmission in affected families.

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References

  1. 1.

    et al. Mitochondrial DNA mutation associated with Leber’s hereditary optic neuropathy. Science 242, 1427–1430 (1988)

  2. 2.

    , & Deletions of muscle mitochondrial DNA in patients with mitochondrial myopathies. Nature 331, 717–719 (1988)

  3. 3.

    et al. Deletions of mitochondrial DNA in Kearns-Sayre syndrome. Neurology 38, 1339–1346 (1988)

  4. 4.

    , , & Genetic diseases of the mitochondrial DNA in humans [in Spanish]. Salud Publica Mex. 43, 151–161 (2001)

  5. 5.

    Mitochondrial dysfunction, oxidative stress, regulation of exocytosis and their relevance to neurodegenerative diseases. J. Neurochem. 104, 298–305 (2008)

  6. 6.

    & Mitochondrial dysfunction and oxidative stress in neurodegenerative diseases. Nature 443, 787–795 (2006)

  7. 7.

    , & Mitochondrial DNA mutations in disease, aging, and neurodegeneration. Ann. NY Acad. Sci. 1147, 21–29 (2008)

  8. 8.

    & Oxidative stress and mitochondrial dysfunction in neurodegenerative diseases. Neuroscience 145, 1233–1248 (2007)

  9. 9.

    , & Mitochondrial mutations in cancer. Oncogene 25, 4647–4662 (2006)

  10. 10.

    et al. Ubiquitin tag for sperm mitochondria. Nature 402, 371–372 (1999)

  11. 11.

    et al. Epidemiology of A3243G, the mutation for mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes: prevalence of the mutation in an adult population. Am. J. Hum. Genet. 63, 447–454 (1998)

  12. 12.

    & Mitochondrial DNA mutations in human disease. Nature Rev. Genet. 6, 389–402 (2005)

  13. 13.

    et al. Prevalence of mitochondrial DNA disease in adults. Ann. Neurol. 63, 35–39 (2008)

  14. 14.

    et al. Analysis of mtDNA variant segregation during early human embryonic development: a tool for successful NARP preimplantation diagnosis. J. Med. Genet. 43, 244–247 (2006)

  15. 15.

    & Mitochondrial disorders: genetics, counseling, prenatal diagnosis and reproductive options. Am. J. Med. Genet. 106, 102–114 (2001)

  16. 16.

    et al. Producing primate embryonic stem cells by somatic cell nuclear transfer. Nature 450, 497–502 (2007)

  17. 17.

    et al. Reprogramming following somatic cell nuclear transfer in primates is dependent upon nuclear remodeling. Hum. Reprod. 22, 2232–2242 (2007)

  18. 18.

    et al. Use of assisted reproductive technologies in the propagation of rhesus macaque offspring. Biol. Reprod. 71, 486–493 (2004)

  19. 19.

    , , & Techniques and pitfalls in the detection of pathogenic mitochondrial DNA mutations. J. Mol. Diagn. 5, 197–208 (2003)

  20. 20.

    et al. Safety evaluation of hemagglutinating virus of Japan—artificial viral envelope liposomes in nonhuman primates. Hum. Gene Ther. 12, 469–487 (2001)

  21. 21.

    , , & Preventing transmission of maternally inherited mitochondrial DNA diseases. BMJ 338, b94 (2009)

  22. 22.

    Prevention of human mitochondrial (mtDNA) disease by nucleus transplantation into an enucleated donor oocyte. Am. J. Med. Genet. 87, 265–266 (1999)

  23. 23.

    et al. Ooplasmic transfer in mature human oocytes. Mol. Hum. Reprod. 4, 269–280 (1998)

  24. 24.

    et al. Pregnancy following transfer of ooplasm from cryopreserved-thawed donor oocytes into recipient oocytes. Fertil. Steril. 71, 575–577 (1999)

  25. 25.

    , , , & Birth of infant after transfer of anucleate donor oocyte cytoplasm into recipient eggs. Lancet 350, 186–187 (1997)

  26. 26.

    , , & Mitochondria in human offspring derived from ooplasmic transplantation. Hum. Reprod. 16, 513–516 (2001)

  27. 27.

    , , & Mitochondrial DNA heteroplasmy after human ooplasmic transplantation. Fertil. Steril. 74, 573–578 (2000)

  28. 28.

    et al. Transmission of mitochondrial DNA disorders: possibilities for the future. Lancet 368, 87–89 (2006)

  29. 29.

    , , , & Effect of treating induced mitochondrial damage on embryonic development and epigenesis. Biol. Reprod. 72, 584–592 (2005)

  30. 30.

    et al. Mitochondrial aggregation patterns and activity in human oocytes and preimplantation embryos. Hum. Reprod. 16, 909–917 (2001)

  31. 31.

    & Mitochondrial genotype segregation in a mouse heteroplasmic lineage produced by embryonic karyoplast transplantation. Genetics 145, 445–451 (1997)

  32. 32.

    & Mitochondrial genotype segregation during preimplantation development in mouse heteroplasmic embryos. Genetics 148, 877–883 (1998)

  33. 33.

    et al. Gene therapy for progeny of mito-mice carrying pathogenic mtDNA by nuclear transplantation. Proc. Natl Acad. Sci. USA 102, 16765–16770 (2005)

  34. 34.

    et al. A reduction of mitochondrial DNA molecules during embryogenesis explains the rapid segregation of genotypes. Nature Genet. 40, 249–254 (2008)

  35. 35.

    , & The mitochondrial DNA genetic bottleneck results from replication of a subpopulation of genomes. Nature Genet. 40, 1484–1488 (2008)

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Acknowledgements

The authors acknowledge the Division of Animal Resources, Surgical Team, Endocrine Technology Core, Imaging & Morphology Core and Molecular & Cellular Biology Core at the Oregon National Primate Research Center for providing expertise and services that contributed to this project. We are grateful to W. Sanger and M. Nelson for karyotyping services, C. Penedo for microsatellite analysis and S. Wong for providing Sendai virus. We thank J. Hennebold, R. Stouffer and D. Wolf for consulting, helpful discussions and critical reading of the manuscript. This study was supported by start-up funds from Oregon National Primate Research Center, Oregon Stem Cell Center and grants from the National Institutes of Health.

Author Contributions S.M. and M.T. conceived the study, designed experiments and conducted ST micromanipulations. M.S. performed ICSI, mitochondrial staining and analysis in oocytes. H.S., J.W. and Y.L. conducted ES cell derivation, characterization and differentiation. M.T., H.M., L.C., H.S. and Y.L. performed DNA/RNA isolations and mtDNA analyses. C.R. and O.K. conducted ovarian stimulations, oocyte recovery, ICSI and embryo transfers. S.M. and M.T. analysed the data and wrote the paper.

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Affiliations

  1. Oregon National Primate Research Center,

    • Masahito Tachibana
    • , Michelle Sparman
    • , Hathaitip Sritanaudomchai
    • , Hong Ma
    • , Lisa Clepper
    • , Joy Woodward
    • , Ying Li
    • , Cathy Ramsey
    • , Olena Kolotushkina
    •  & Shoukhrat Mitalipov
  2. Oregon Stem Cell Center and,

    • Shoukhrat Mitalipov
  3. Departments of Obstetrics and Gynecology and Molecular and Medical Genetics, Oregon Health and Science University, 505 N.W. 185th Avenue, Beaverton, Oregon 97006, USA

    • Shoukhrat Mitalipov

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Corresponding author

Correspondence to Shoukhrat Mitalipov.

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    Supplementary Information

    This file contains Supplementary Figures 1-6 with Legends, Supplementary Tables 1-5, Supplementary Methods and Supplementary References.

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DOI

https://doi.org/10.1038/nature08368

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