Targeting specificity of APOBEC-based cytosine base editor in human iPSCs determined by whole genome sequencing

Nat Commun. 2019 Nov 25;10(1):5353. doi: 10.1038/s41467-019-13342-8.

Abstract

DNA base editors have enabled genome editing without generating DNA double strand breaks. The applications of this technology have been reported in a variety of animal and plant systems, however, their editing specificity in human stem cells has not been studied by unbiased genome-wide analysis. Here we investigate the fidelity of cytidine deaminase-mediated base editing in human induced pluripotent stem cells (iPSCs) by whole genome sequencing after sustained or transient base editor expression. While base-edited iPSC clones without significant off-target modifications are identified, this study also reveals the potential of APOBEC-based base editors in inducing unintended point mutations outside of likely in silico-predicted CRISPR-Cas9 off-targets. The majority of the off-target mutations are C:G->T:A transitions or C:G->G:C transversions enriched for the APOBEC mutagenesis signature. These results demonstrate that cytosine base editor-mediated editing may result in unintended genetic modifications with distinct patterns from that of the conventional CRISPR-Cas nucleases.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • APOBEC Deaminases / genetics
  • APOBEC Deaminases / metabolism*
  • Animals
  • CRISPR-Cas Systems
  • Cytidine Deaminase / genetics
  • Cytidine Deaminase / metabolism*
  • Cytosine / metabolism*
  • DNA Breaks, Double-Stranded
  • Gene Editing / methods*
  • Genome, Human / genetics
  • Humans
  • Induced Pluripotent Stem Cells / metabolism*
  • Mutation
  • Plant Cells / metabolism
  • Reproducibility of Results
  • Whole Genome Sequencing / methods*

Substances

  • Cytosine
  • APOBEC Deaminases
  • Cytidine Deaminase