Dissecting Immune Circuits by Linking CRISPR-Pooled Screens with Single-Cell RNA-Seq
Dissecting Immune Circuits by Linking CRISPR-Pooled Screens with Single-Cell RNA-Seq
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DOI：
10.1016/j.cell.2016.11.039
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发表时间：
2016-12-15
期刊：
影响因子：
64.5
通讯作者：
Amit, Ido
中科院分区：
文献类型：
--
作者：
Jaitin, Diego Adhemar;Weiner, Assaf;Amit, Ido
在多细胞生物中，专用的调节电路控制细胞类型的多样性和反应。这些电路中的串扰和冗余和大量的细胞异质性提出了重大的研究挑战。在这里，我们提出了CrispSeq，这是一种用于大规模平行的单细胞RNA测序（RNA-SEQ）的集成方法，并群集定期插入了短期间隔短的腔膜重复序列（CRISPR）pooled屏幕。我们表明，同一细胞中的基因组扰动和转录组的分析使我们能够同时阐明多种因素及其相互作用的功能。我们将Crisp-Seq应用于先天免疫的探测监管电路。通过在体外和小鼠中对成千上万的扰动细胞取样，我们确定了发育依赖性因素和信号依赖性因素之间的相互作用和冗余。其中包括CEBPB和IRF8在调节单核细胞/巨噬细胞与树突状细胞谱系中的相反作用，以及RELA和STAT1/2在单核细胞中的差异功能以及对病原体的树突状细胞反应。这项研究确立了Crisp-Seq是一种阐明哺乳动物调节电路的广泛，全面和无偏见的方法。
In multicellular organisms, dedicated regulatory circuits control cell type diversity and responses. The crosstalk and redundancies within these circuits and substantial cellular heterogeneity pose a major research challenge. Here, we present CRISPseq, an integrated method for massively parallel single-cell RNA sequencing (RNA-seq) and clustered regularly interspaced short palindromic repeats (CRISPR)-pooled screens. We show that profiling the genomic perturbation and transcriptome in the same cell enables us to simultaneously elucidate the function of multiple factors and their interactions. We applied CRISP-seq to probe regulatory circuits of innate immunity. By sampling tens of thousands of perturbed cells in vitro and in mice, we identified interactions and redundancies between developmental and signaling-dependent factors. These include opposing effects of Cebpb and Irf8 in regulating the monocyte/macrophage versus dendritic cell lineages and differential functions for Rela and Stat1/2 in monocyte versus dendritic cell responses to pathogens. This study establishes CRISP-seq as a broadly applicable, comprehensive, and unbiased approach for elucidating mammalian regulatory circuits.