Genome-wide Cas9-mediated screening of essential non-coding regulatory elements via libraries of paired single-guide RNAs
Nature Biomedical Engineering, vol. 8, pp. 890–908
Abstract
The functions of non-coding regulatory elements (NCREs), which constitute a major fraction of the human genome, have not been systematically studied. Here we report a method involving libraries of paired single-guide RNAs targeting both ends of an NCRE as a screening system for the Cas9-mediated deletion of thousands of NCREs genome-wide to study their functions in distinct biological contexts. By using K562 and 293T cell lines and human embryonic stem cells, we show that NCREs can have redundant functions, and that many ultra-conserved elements have silencer activity and play essential roles in cell growth and in cellular responses to drugs (notably, the ultra-conserved element PAX6_Tarzan may be critical for heart development, as removing it from human embryonic stem cells led to defects in cardiomyocyte differentiation). The high-throughput screen, which is compatible with single-cell sequencing, may allow for the identification of druggable NCREs.
Authors 15
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Leiden University Medical Center
Affiliation as printed
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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Leiden University Medical Center
Affiliation as printed
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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Leiden University Medical Center
Affiliation as printed
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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Leiden University Medical Center
Affiliation as printed
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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Leiden University Medical Center
Affiliation as printed
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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Leiden University Medical Center
Affiliation as printed
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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Leiden University Medical Center
Affiliation as printed
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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Leiden University Medical Center
Affiliation as printed
Leiden Genome Technology Center, Department of Human Genetics, Leiden University Medical Center, Leiden, the Netherlands
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Leiden University Medical Center
Affiliation as printed
Leiden Genome Technology Center, Department of Human Genetics, Leiden University Medical Center, Leiden, the Netherlands
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Richard P. Davis Aachen Department of Anatomy and Embryology Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW)
Leiden University Medical Center · Novo Nordisk Foundation Center for Stem Cell Medicine
Affiliation as printed
Department of Anatomy and Embryology, The Novo Nordisk Foundation Center for Stem Cell Medicine (reNEW), Leiden University Medical Center, Leiden, the Netherlands
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Leiden University Medical Center
Affiliation as printed
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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The University of Texas Southwestern Medical Center
Affiliation as printed
Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, TX, USA
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Michael Snyder corresponding
Affiliation as printed
Department of Genetics, Stanford University, Stanford, CA, USA. mpsnyder@stanford.edu
Department of Genetics, Stanford University, Stanford, CA, USA
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Xiao Li corresponding
Case Western Reserve University
Affiliation as printed
Department of Biochemistry, The Center for RNA Science and Therapeutics, Department of Computer and Data Sciences, Case Western Reserve University, Cleveland, OH, USA. xiao.li9@case.edu
Department of Biochemistry, The Center for RNA Science and Therapeutics, Department of Computer and Data Sciences, Case Western Reserve University, Cleveland, OH, USA
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Leiden University Medical Center
Affiliation as printed
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands. b.pang@lumc.nl
Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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