De novo identification of essential protein domains from CRISPR-Cas9 tiling-sgRNA knockout screens
De novo identification of essential protein domains from CRISPR-Cas9 tiling-sgRNA knockout screens
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DOI：
10.1038/s41467-019-12489-8
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发表时间：
2019-10-04
影响因子：
16.6
通讯作者：
Xu, Han
中科院分区：
文献类型：
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作者：
He, Wei;Zhang, Liang;Xu, Han
使用平铺sgRNA设计的高通量CRISPR-Cas9敲除筛选允许原位评估蛋白质结构域功能。在这里，为了便于从这样的筛选中从头鉴定必需的蛋白质结构域，我们提出了ProTiler，这是一种用于CRISPR敲除超敏感（CKHS）区域的稳健映射的计算方法，CKHS区域是指与筛选中的强sgRNA缺失效应相关的蛋白质区域。应用于已发表的CRISPR平铺屏幕数据集，ProTiler在83种蛋白质中识别出175个CKHS区域。在这些CKHS区域中，超过80%与注释的Pfam结构域重叠，包括数据集中所有15个已知的药物靶点。ProTiler还揭示了未注释的必需结构域，包括SWI/SNF亚基SMARCB 1的N-末端，这是实验验证的。令人惊讶的是，CKHS区域与磷酸化和乙酰化位点负相关，表明蛋白质结构域和翻译后修饰位点对CRISPR-Cas9介导的氨基酸丢失具有不同的敏感性。
High-throughput CRISPR-Cas9 knockout screens using a tiling-sgRNA design permit in situ evaluation of protein domain function. Here, to facilitate de novo identification of essential protein domains from such screens, we propose ProTiler, a computational method for the robust mapping of CRISPR knockout hyper-sensitive (CKHS) regions, which refer to the protein regions associated with a strong sgRNA dropout effect in the screens. Applied to a published CRISPR tiling screen dataset, ProTiler identifies 175 CKHS regions in 83 proteins. Of these CKHS regions, more than 80% overlap with annotated Pfam domains, including all of the 15 known drug targets in the dataset. ProTiler also reveals unannotated essential domains, including the N-terminus of the SWI/SNF subunit SMARCB1, which is validated experimentally. Surprisingly, the CKHS regions are negatively correlated with phosphorylation and acetylation sites, suggesting that protein domains and post-translational modification sites have distinct sensitivities to CRISPR-Cas9 mediated amino acids loss.