Repair of DNA double-strand breaks by templated nucleotide sequence insertions derived from distant regions of the genome

Masahiro Onozawa, Zhenhua Zhang, Yoo Jung Kim, Liat Goldberg, Tamas Varga, P. Leif Bergsagel, W. Michael Kuehl, Peter D. Aplan

Research output: Contribution to journalArticle

37 Scopus citations

Abstract

We used the I-SceI endonuclease to produce DNA double-strand breaks (DSBs) and observed that a fraction of these DSBs were repaired by insertion of sequences, which we termed "templated sequence insertions" (TSIs), derived from distant regions of the genome. These TSIs were derived from genic, retrotransposon, or telomere sequences and were not deleted from the donor site in the genome, leading to the hypothesis that they were derived from reverse-transcribed RNA. Cotransfection of RNA and an I-SceI expression vector demonstrated insertion of RNA-derived sequences at the DNA-DSB site, and TSIs were suppressed by reverse-transcriptase inhibitors. Both observations support the hypothesis that TSIs were derived from RNA templates. In addition, similar insertions were detected at sites of DNA DSBs induced by transcription activator-like effector nuclease proteins. Whole-genome sequencing of myeloma cell lines revealed additional TSIs, demonstrating that repair of DNA DSBs via insertion was not restricted to experimentally produced DNA DSBs. Analysis of publicly available databases revealed that many of these TSIs are polymorphic in the human genome. Taken together, these results indicate that inser-tional events should be considered as alternatives to gross chromosomal rearrangements in the interpretation of whole-genome sequence data and that this mutagenic form of DNA repair may play a role in genetic disease, exon shuffling, and mammalian evolution.

Original languageEnglish (US)
Pages (from-to)7729-7734
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume111
Issue number21
DOIs
StatePublished - May 27 2014

Keywords

  • DNA patch
  • LINE-1
  • Polymorphism
  • TSIP

ASJC Scopus subject areas

  • General

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