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Cell. 2016 Dec 15;167(7):1883-1896.e15. doi: 10.1016/j.cell.2016.11.039.

Dissecting Immune Circuits by Linking CRISPR-Pooled Screens with Single-Cell RNA-Seq.

Author information

1
Department of Immunology, Weizmann Institute, Rehovot 76100, Israel.
2
Department of Immunology, Weizmann Institute, Rehovot 76100, Israel; Hubrecht Institute-KNAW, Royal Netherlands Academy of Arts and Sciences, 3584 CT Utrecht, the Netherlands.
3
Flow Cytometry Unit, Department of Biological Services, Weizmann Institute of Science, Rehovot 76100, Israel.
4
Department of Computer Science and Applied Mathematics, Department of Biological Regulation, Weizmann Institute, Rehovot 76100, Israel.
5
Hubrecht Institute-KNAW, Royal Netherlands Academy of Arts and Sciences, 3584 CT Utrecht, the Netherlands.
6
Department of Immunology, Weizmann Institute, Rehovot 76100, Israel. Electronic address: ido.amit@weizmann.ac.il.

Abstract

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 CRISP-seq, 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.

KEYWORDS:

CRISPR/Cas9; RNA-seq; functional genomics; gene networks; genetic screen; immune response; innate immunity; single-cell RNA-seq; transcriptomics

Comment in

PMID:
27984734
DOI:
10.1016/j.cell.2016.11.039
[Indexed for MEDLINE]
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