Genomic Copy Number Dictates a Gene-Independent Cell Response to CRISPR/Cas9 Targeting.
Genomic Copy Number Dictates a Gene-Independent Cell Response to CRISPR/Cas9 Targeting.
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DOI：
10.1158/2159-8290.cd-16-0154
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发表时间：
2016-08
期刊：
影响因子：
28.2
通讯作者：
Hahn WC
中科院分区：
文献类型：
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作者：
Aguirre AJ;Meyers RM;Weir BA;Vazquez F;Zhang CZ;Ben-David U;Cook A;Ha G;Harrington WF;Doshi MB;Kost-Alimova M;Gill S;Xu H;Ali LD;Jiang G;Pantel S;Lee Y;Goodale A;Cherniack AD;Oh C;Kryukov G;Cowley GS;Garraway LA;Stegmaier K;Roberts CW;Golub TR;Meyerson M;Root DE;Tsherniak A;Hahn WC
CRISPR-Cas9 系统能够在哺乳动物细胞中进行基因组编辑和体细胞遗传筛选。我们在 33 种癌细胞系中进行了基因组规模的功能丧失筛查，以确定增殖/生存必需的基因，并发现基因组编辑后基因拷贝数增加与细胞活力下降之间存在很强的相关性。在拷贝数增加的区域内，CRISPR-Cas9 靶向表达和未表达的基因以及基因间位点，通过诱导 G2 细胞周期停滞，导致细胞增殖显着降低。通过检查映射到多个基因组位点的单引导 RNA，我们发现细胞对 CRISPR-Cas9 编辑的反应与目标基因座的数量密切相关。这些观察结果表明，CRISPR-Cas9 的基因组靶向引发了不依赖于基因的抗增殖细胞反应。这种效应对于解释 CRISPR-Cas9 筛选数据具有重要的实际意义，并且混淆了使用该技术识别扩增区域中的必需基因。
The CRISPR-Cas9 system enables genome editing and somatic cell genetic screens in mammalian cells. We performed genome scale loss-of-function screens in 33 cancer cell lines to identify genes essential for proliferation/survival and found a strong correlation between increased gene copy number and decreased cell viability after genome editing. Within regions of copy number gain, CRISPR-Cas9 targeting of both expressed and unexpressed genes, as well as intergenic loci, led to significantly decreased cell proliferation through induction of a G2 cell cycle arrest. By examining single guide RNAs that map to multiple genomic sites, we found that this cell response to CRISPR-Cas9 editing correlated strongly with the number of target loci. These observations indicate that genome targeting by CRISPR-Cas9 elicits a gene-independent anti-proliferative cell response. This effect has important practical implications for interpretation of CRISPR-Cas9 screening data and confounds the use of this technology for identification of essential genes in amplified regions.