Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation
- PMID: 25307932
- PMCID: PMC4253859
- DOI: 10.1016/j.cell.2014.09.029
Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation
Abstract
While the catalog of mammalian transcripts and their expression levels in different cell types and disease states is rapidly expanding, our understanding of transcript function lags behind. We present a robust technology enabling systematic investigation of the cellular consequences of repressing or inducing individual transcripts. We identify rules for specific targeting of transcriptional repressors (CRISPRi), typically achieving 90%-99% knockdown with minimal off-target effects, and activators (CRISPRa) to endogenous genes via endonuclease-deficient Cas9. Together they enable modulation of gene expression over a ∼1,000-fold range. Using these rules, we construct genome-scale CRISPRi and CRISPRa libraries, each of which we validate with two pooled screens. Growth-based screens identify essential genes, tumor suppressors, and regulators of differentiation. Screens for sensitivity to a cholera-diphtheria toxin provide broad insights into the mechanisms of pathogen entry, retrotranslocation and toxicity. Our results establish CRISPRi and CRISPRa as powerful tools that provide rich and complementary information for mapping complex pathways.
Copyright © 2014 Elsevier Inc. All rights reserved.
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Comment in
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Genetic screens: CRISPR screening from both ways.Nat Rev Genet. 2014 Dec;15(12):778-9. doi: 10.1038/nrg3850. Epub 2014 Oct 21. Nat Rev Genet. 2014. PMID: 25331481 No abstract available.
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Toward Whole-Transcriptome Editing with CRISPR-Cas9.Mol Cell. 2015 May 21;58(4):560-2. doi: 10.1016/j.molcel.2015.05.016. Mol Cell. 2015. PMID: 26000839
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