Eya2 promotes cell cycle progression by regulating DNA damage response during vertebrate limb regeneration

  1. Konstantinos Sousounis
  2. Donald M Bryant
  3. Jose Martinez Fernandez
  4. Samuel S Eddy
  5. Stephanie L Tsai
  6. Gregory C Gundberg
  7. Jihee Han
  8. Katharine Courtemanche
  9. Michael Levin
  10. Jessica L Whited  Is a corresponding author
  1. Harvard University, United States
  2. Brigham & Women's Hospital, United States
  3. Tufts University, United States
  4. Harvard Medical School, United States

Abstract

How salamanders accomplish progenitor cell proliferation while faithfully maintaining genomic integrity and regenerative potential remains elusive. Here we found an innate DNA damage response mechanism that is evident during blastema proliferation (early- to late-bud) and studied its role during tissue regeneration by ablating the function of one of its components, Eyes absent 2. In eya2 mutant axolotls, we found that DNA damage signaling through the H2AX histone variant was deregulated, especially within the proliferating progenitors during limb regeneration. Ultimately, cell cycle progression was impaired at the G1/S and G2/M transitions and regeneration rate was reduced. Similar data were acquired using acute pharmacological inhibition of the Eya2 phosphatase activity and the DNA damage checkpoint kinases Chk1 and Chk2 in wild-type axolotls. Together, our data indicate that highly-regenerative animals employ a robust DNA damage response pathway which involves regulation of H2AX phosphorylation via Eya2 to facilitate proper cell cycle progression upon injury.

Data availability

Raw data can be accessed in the NIH Sequence Read Archive: SUB6297224.

The following previously published data sets were used

Article and author information

Author details

  1. Konstantinos Sousounis

    Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Donald M Bryant

    Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Jose Martinez Fernandez

    Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Samuel S Eddy

    Orthopedic Surgery, Brigham & Women's Hospital, Boston, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Stephanie L Tsai

    Stem Cell and Regenerative Biology, Molecular and Cellular Biology, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7549-3418
  6. Gregory C Gundberg

    Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Jihee Han

    Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Katharine Courtemanche

    Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
  9. Michael Levin

    The Allen Discovery Center, Tufts University, Medford, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7292-8084
  10. Jessica L Whited

    Department of Orthopedic Surgery, Harvard Medical School, Boston, United States
    For correspondence
    jwhited@partners.org
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3709-6515

Funding

Sara Elizabeth O'Brien Trust (Postdoctoral fellowship)

  • Konstantinos Sousounis

National Institutes of Health (K99EY029361)

  • Konstantinos Sousounis

Paul G. Allen Family Foundation (Allen Discovery Center at Tufts)

  • Michael Levin
  • Jessica L Whited

National Institutes of Health (1DP2HD087953)

  • Jessica L Whited

National Institutes of Health (1R01HD095494)

  • Jessica L Whited

Harvard Stem Cell Institute (HIP)

  • Jose Martinez Fernandez

Howard Hughes Medical Institute (Gilliam Fellowship)

  • Donald M Bryant

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: All experiments involving animals were performed according to IACUC protocol #2016N000369 at Brigham and Women's Hospital. All surgeries were performed while animals were anesthetized in tricaine. All experiments were planned and executed in manners that minimized animal suffering.

Reviewing Editor

  1. Stephen Randal Voss, University of Kentucky

Version history

  1. Received: August 20, 2019
  2. Accepted: March 5, 2020
  3. Accepted Manuscript published: March 6, 2020 (version 1)
  4. Version of Record published: March 24, 2020 (version 2)

Copyright

© 2020, Sousounis et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Konstantinos Sousounis
  2. Donald M Bryant
  3. Jose Martinez Fernandez
  4. Samuel S Eddy
  5. Stephanie L Tsai
  6. Gregory C Gundberg
  7. Jihee Han
  8. Katharine Courtemanche
  9. Michael Levin
  10. Jessica L Whited
(2020)
Eya2 promotes cell cycle progression by regulating DNA damage response during vertebrate limb regeneration
eLife 9:e51217.
https://doi.org/10.7554/eLife.51217

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