Genome-wide microhomologies enable precise template-free editing of biologically relevant deletion mutations

Janin Grajcarek, Jean Monlong, Yoko Nishinaka-Arai, Michiko Nakamura, Miki Nagai, Shiori Matsuo, David Lougheed, Hidetoshi Sakurai, Megumu K. Saito, Guillaume Bourque and Knut Woltjen ()
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Janin Grajcarek: Kyoto University
Jean Monlong: McGill University
Yoko Nishinaka-Arai: Kyoto University
Michiko Nakamura: Kyoto University
Miki Nagai: Kyoto University
Shiori Matsuo: Kyoto University
David Lougheed: Canadian Center for Computational Genomics
Hidetoshi Sakurai: Kyoto University
Megumu K. Saito: Kyoto University
Guillaume Bourque: McGill University
Knut Woltjen: Kyoto University

Nature Communications, 2019, vol. 10, issue 1, 1-13

Abstract: Abstract The functional effect of a gene edit by designer nucleases depends on the DNA repair outcome at the targeted locus. While non-homologous end joining (NHEJ) repair results in various mutations, microhomology-mediated end joining (MMEJ) creates precise deletions based on the alignment of flanking microhomologies (µHs). Recently, the sequence context surrounding nuclease-induced double strand breaks (DSBs) has been shown to predict repair outcomes, for which µH plays an important role. Here, we survey naturally occurring human deletion variants and identify that 11 million or 57% are flanked by µHs, covering 88% of protein-coding genes. These biologically relevant mutations are candidates for precise creation in a template-free manner by MMEJ repair. Using CRISPR-Cas9 in human induced pluripotent stem cells (hiPSCs), we efficiently create pathogenic deletion mutations for demonstrable disease models with both gain- and loss-of-function phenotypes. We anticipate this dataset and gene editing strategy to enable functional genetic studies and drug screening.

Date: 2019
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DOI: 10.1038/s41467-019-12829-8

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