The structure-selective endonucleases GEN1 and MUS81 mediate complementary functions in safeguarding the genome of proliferating B lymphocytes

  1. Keith Conrad Fernandez
  2. Laura Feeney
  3. Ryan M Smolkin
  4. Wei-Feng Yen
  5. Allysia J Matthews
  6. John HJ Petrini  Is a corresponding author
  7. Jayanta Chaudhuri  Is a corresponding author
  1. Memorial Sloan Kettering Cancer Center, United States

Abstract

During the development of humoral immunity, activated B lymphocytes undergo vigorous proliferative, transcriptional, metabolic, and DNA remodeling activities; hence, their genomes are constantly exposed to an onslaught of genotoxic agents and processes. Branched DNA intermediates generated during replication and recombinational repair must be eliminated to preserve the integrity of these DNA transactions for the faithful duplication and propagation of genomic material. To investigate the role of two structure-selective endonucleases, GEN1 and MUS81, in B cell biology, we established B-cell conditional knockout mouse models and found that targeted deletion of GEN1 and MUS81 in early B-cell precursors abrogates the development and maturation of B-lineage cells while selective loss of the enzymes in mature B cells inhibits the generation of robust germinal centers. Upon activation, these double-null mature B lymphocytes fail to proliferate and survive while exhibiting transcriptional signatures of p53 signaling, apoptosis, and type I interferon response. Metaphase spreads of these endonuclease-deficient cells showed severe and diverse chromosomal abnormalities, including a preponderance of chromosome breaks, consistent with a defect in resolving DNA recombination intermediates. These observations underscore the pivotal roles of GEN1 and MUS81 in safeguarding the genome to ensure the proper development and maintenance of B lymphocytes.

Data availability

The RNAseq data and analysis generated in this study are deposited in GEO under the accession code GSE195734. The Gen1 and Mus81 expression data in the various B cell subsets was previously generated by Brazão et al. (2016) and deposited under GSE72018 in GEO.

The following data sets were generated
The following previously published data sets were used

Article and author information

Author details

  1. Keith Conrad Fernandez

    Immunology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Laura Feeney

    Molecular Biology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Ryan M Smolkin

    Immunology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Wei-Feng Yen

    Immunology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Allysia J Matthews

    Immunology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. John HJ Petrini

    Molecular Biology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    For correspondence
    petrinij@mskcc.org
    Competing interests
    The authors declare that no competing interests exist.
  7. Jayanta Chaudhuri

    Immunology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    For correspondence
    chaudhuj@mskcc.org
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3838-0075

Funding

National Institutes of Health (R01AI072194)

  • Jayanta Chaudhuri

National Institutes of Health (R01AI124186)

  • Jayanta Chaudhuri

National Institutes of Health (R56AI072194)

  • Jayanta Chaudhuri

National Institutes of Health (U54CA137788)

  • Jayanta Chaudhuri

National Institutes of Health (P30CA008748)

  • John HJ Petrini
  • Jayanta Chaudhuri

National Institutes of Health (R01GM56888)

  • John HJ Petrini

National Institutes of Health (R35GM136278)

  • John HJ Petrini

National Institutes of Health (U54OD020355)

  • John HJ Petrini

Geoffrey Beene Cancer Research Center

  • Jayanta Chaudhuri

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 mice were housed and maintained in groups of five under specific pathogen-free conditions, and euthanized at the time of analyses in accordance with guidelines for animal care established by Memorial Sloan Kettering Cancer Center Research Animal Resource Center and the Institutional Animal Care and Use Committee (IACUC). All mouse experimentation protocols were approved by MSK's IACUC (Protocol Number: 05-12-030).

Reviewing Editor

  1. Wolf-Dietrich Heyer, University of California, Davis, United States

Publication history

  1. Received: January 14, 2022
  2. Preprint posted: February 10, 2022 (view preprint)
  3. Accepted: September 30, 2022
  4. Accepted Manuscript published: October 3, 2022 (version 1)
  5. Version of Record published: October 19, 2022 (version 2)
  6. Version of Record updated: October 26, 2022 (version 3)

Copyright

© 2022, Fernandez 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. Keith Conrad Fernandez
  2. Laura Feeney
  3. Ryan M Smolkin
  4. Wei-Feng Yen
  5. Allysia J Matthews
  6. John HJ Petrini
  7. Jayanta Chaudhuri
(2022)
The structure-selective endonucleases GEN1 and MUS81 mediate complementary functions in safeguarding the genome of proliferating B lymphocytes
eLife 11:e77073.
https://doi.org/10.7554/eLife.77073

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