A Genome-wide Framework for Mapping Gene Regulation via Cellular Genetic Screens
A Genome-wide Framework for Mapping Gene Regulation via Cellular Genetic Screens
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DOI：
10.1016/j.cell.2018.11.029
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发表时间：
2019-01-10
期刊：
影响因子：
64.5
通讯作者：
Shendure, Jay
中科院分区：
文献类型：
--
作者：
Gasperini, Molly;Hill, Andrew J.;Shendure, Jay
在人类基因组中已经确定了超过100万个候选调控元件，但几乎所有的调控元件都是未经验证的，它们的靶基因也不确定。基于人类遗传学的方法仅限于常见的变异和通过连锁不平衡来解决问题。我们提出了一个多重、表达数量性状位点（eQTL）启发的框架，通过将CRISPR/ cas9介导的扰动随机组合引入许多细胞中的每个细胞，然后进行单细胞RNA测序（RNA-seq），来定位增强基因对。在两个实验中，我们使用dCas9-KRAB干扰5,920个候选增强子，对其靶基因没有很强的先验假设，通过分析254,974个单细胞转录组来测量效果。我们确定了664对（470对高置信度）顺式增强基因对，这些基因对富集了特定的转录因子，非管家状态，基因组和3D构象接近其靶基因。这个框架将促进增强基因调控相互作用的大规模定位，这是人类基因组顺式调控景观的一个关键但很大程度上未知的组成部分。
Over one million candidate regulatory elements have been identified across the human genome, but nearly all are unvalidated and their target genes uncertain. Approaches based on human genetics are limited in scope to common variants and in resolution by linkage disequilibriunn. We present a multiplex, expression quantitative trait locus (eQTL)-inspired framework for mapping enhancer-gene pairs by introducing random combinations of CRISPR/Cas9-mediated perturbations to each of many cells, followed by single-cell RNA sequencing (RNA-seq). Across two experiments, we used dCas9-KRAB to perturb 5,920 candidate enhancers with no strong a priori hypothesis as to their target gene(s), measuring effects by profiling 254,974 single cell transcriptomes. We identified 664 (470 high confidence) cis enhancer-gene pairs, which were enriched for specific transcription factors, non housekeeping status, and genomic and 3D conformational proximity to their target genes. This framework will facilitate the large-scale mapping of enhancer-gene regulatory interactions, a critical yet largely uncharted component of the cis-regulatory landscape of the human genome.