Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation
Name
Ploegh_Genome-scale CRISPER.pdf
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1.66 MB
Format
Adobe PDF
Checksum (MD5)
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Author(s)
Gilbert, Luke A.
Horlbeck, Max A.
Adamson, Britt
Villalta, Jacqueline E.
Chen, Yuwen
Whitehead, Evan H.
Guimaraes, Carla
Panning, Barbara
Bassik, Michael C.
Qi, Lei S.
Date Issued
October 2014
Journal
Cell
Publisher
Elsevier
Citation
Gilbert, Luke A. et al. “Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation.” Cell 159.3 (2014): 647–661.
Version
Author's final manuscript
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.
MIT Department
Massachusetts Institute of Technology. Department of Biology
Whitehead Institute for Biomedical Research
Terms of Use
Creative Commons Attribution-NonCommercial-NoDerivs License
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DOI of Published Version
https://doi.org/10.1016/j.cell.2014.09.029
