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APOBEC3 induces mutations during repair of CRISPR-Cas9-generated DNA breaks

Liqun Lei1,2,3, Hongquan Chen4,5, Wei Xue6

  • 1School of Life Science and Technology, ShanghaiTech University, Shanghai, China.

Nature Structural & Molecular Biology
|January 12, 2018
PubMed

Insights

APOBEC3 enzymes cause mutations in DNA repaired after CRISPR-Cas9 editing. Suppressing APOBEC3 or using specific repair methods can reduce these unwanted genomic DNA mutations.

Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • The APOBEC-AID family of cytidine deaminases targets single-stranded nucleic acids.
  • Single-stranded DNA is involved in DNA repair pathways, including those for CRISPR-Cas9-induced breaks.

Purpose of the Study:

  • To investigate the role of APOBEC3 in generating mutations during CRISPR-Cas9 DNA repair in human cells.
  • To identify strategies for minimizing unwanted mutations in genomic DNA after CRISPR-Cas9 editing.

Main Methods:

  • Studied APOBEC3 activity on single-stranded oligodeoxynucleotides in human cells.
  • Analyzed mutations resulting from homology-directed repair (HDR) of Cas9-generated double-strand breaks.
  • Investigated APOBEC3-mediated deamination during repair of Cas9 nickase-generated single-strand breaks.

Main Results:

  • APOBEC3 triggers cytidine deamination of single-stranded oligodeoxynucleotides, leading to base substitutions via HDR of Cas9-induced double-strand breaks.
  • APOBEC3-catalyzed deamination in single-stranded DNA during repair of Cas9 nickase breaks results in insertions or deletions (indels).
  • Both APOBEC3 and DNA repair proteins contribute to indel generation.

Conclusions:

  • Optimizing CRISPR-Cas9 repair conditions, such as using double-stranded donors for HDR or suppressing APOBEC3, can reduce unwanted genomic mutations.
  • Understanding APOBEC3's role is crucial for precise genome editing applications.

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