Optical Pooled Screens in Human Cells

被引：174

作者：

Feldman, David ^{[1
,2
]}

Singh, Avtar ^{[1
]}

Schmid-Burgk, Jonathan L. ^{[1
]}

Carlson, Rebecca J. ^{[1
,3
]}

Mezger, Anja ^{[1
,4
]}

Garrity, Anthony J. ^{[1
]}

Zhang, Feng ^{[1
,5
,6
,7
,8
]}

Blainey, Paul C. ^{[1
,5
,9
]}

机构：

[1] Broad Inst MIT & Harvard, Cambridge, MA 02142 USA

[2] MIT, Dept Phys, Cambridge, MA 02142 USA

[3] MIT, Dept Hlth Sci & Technol, Cambridge, MA 02142 USA

[4] Karolinska Inst, Dept Med Biochem & Biophys, Stockholm, Sweden

[5] MIT, Dept Biol Engn, Cambridge, MA 02142 USA

[6] MIT, McGovern Inst Brain Res, Cambridge, MA 02142 USA

[7] MIT, Dept Brain & Cognit Sci, Cambridge, MA 02142 USA

[8] MIT, Howard Hughes Med Inst, Cambridge, MA 02142 USA

[9] MIT, Koch Inst Integrat Canc Res, Cambridge, MA 02142 USA

来源：

CELL | 2019年 / 179卷 / 03期

基金：

瑞典研究理事会;

关键词：

NF-KAPPA-B; WIDE SIRNA SCREEN; SYSTEMATIC DISSECTION; GENOME; GENES; ACTIVATION; FAMILY; TNF; TRANSCRIPTION; CIRCUITS;

D O I：

10.1016/j.cell.2019.09.016

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

Genetic screens are critical for the systematic identification of genes underlying cellular phenotypes. Pooling gene perturbations greatly improves scalability but is not compatible with imaging of complex and dynamic cellular phenotypes. Here, we introduce a pooled approach for optical genetic screens in mammalian cells. We use targeted in situ sequencing to demultiplex a library of genetic perturbations following image-based phenotyping. We screened a set of 952 genes across millions of cells for involvement in nuclear factor kB (NF-kappa B) signaling by imaging the translocation of RelA (p65) to the nucleus. Screening at a single time point across 3 cell lines recovered 15 known pathway components, while repeating the screen with live-cell imaging revealed a role for Mediator complex subunits in regulating the duration of p65 nuclear retention. These results establish a highly multiplexed approach to imagebased screens of spatially and temporally defined phenotypes with pooled libraries.

引用

页码：787 / +

页数：30

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