Genome-scale CRISPR screens are efficient in non-homologous end-joining deficient cells
Genome-scale CRISPR screens are efficient in non-homologous end-joining deficient cells
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DOI：
10.1038/s41598-019-52078-9
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发表时间：
2019-10-31
影响因子：
4.6
通讯作者：
Loizou, Joanna I.
中科院分区：
文献类型：
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作者：
da Silva, Joana Ferreira;Salic, Sejla;Loizou, Joanna I.
Cas9产生的断裂的诱变修复被认为主要依赖于非同源末端连接（NHEJ），导致DNA内的插入和缺失，最终导致基因敲除（KO）。在这项研究中，通过采取集中以及全基因组的方法，我们表明，这条途径是修复这种病变的关键。NHEJ的基因消融通过替代性末端连接（alt-EJ）以POLQ依赖性方式完全补偿，从而产生具有较大缺失的独特修复特征，其可用于大规模基因组编辑。此外，我们还发现，缺乏NHEJ和alt-EJ的细胞仍然能够修复CRISPR介导的DNA双链断裂，这凸显了人们对基于CRISPR的基因组编辑机制知之甚少。
The mutagenic repair of Cas9 generated breaks is thought to predominantly rely on non-homologous end-joining (NHEJ), leading to insertions and deletions within DNA that culminate in gene knock-out (KO). In this study, by taking focused as well as genome-wide approaches, we show that this pathway is dispensable for the repair of such lesions. Genetic ablation of NHEJ is fully compensated for by alternative end joining (alt-EJ), in a POLQ-dependent manner, resulting in a distinct repair signature with larger deletions that may be exploited for large-scale genome editing. Moreover, we show that cells deficient for both NHEJ and alt-EJ were still able to repair CRISPR-mediated DNA double-strand breaks, highlighting how little is yet known about the mechanisms of CRISPR-based genome editing.