Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complex
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Author(s)
Konermann, Silvana
Brigham, Mark D.
Trevino, Alexandro E.
Joung, Julia
Barcena, Clea
Hsu, Patrick D.
Habib, Naomi
Gootenberg, Jonathan S.
Nishimasu, Hiroshi
Nureki, Osamu
Date Issued
May 22, 2016
Journal
Nature
Citation
Konermann, Silvana, Mark D. Brigham, Alexandro E. Trevino, Julia Joung, Omar O. Abudayyeh, Clea Barcena, Patrick D. Hsu, et al. “Genome-Scale Transcriptional Activation by an Engineered CRISPR-Cas9 Complex.” Nature 517, no. 7536 (December 10, 2014): 583–88.
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Author's final manuscript
Abstract
Systematic interrogation of gene function requires the ability to perturb gene expression in a robust and generalizable manner. Here we describe structure-guided engineering of a CRISPR-Cas9 complex to mediate efficient transcriptional activation at endogenous genomic loci. We used these engineered Cas9 activation complexes to investigate single-guide RNA (sgRNA) targeting rules for effective transcriptional activation, to demonstrate multiplexed activation of ten genes simultaneously, and to upregulate long intergenic non-coding RNA (lincRNA) transcripts. We also synthesized a library consisting of 70,290 guides targeting all human RefSeq coding isoforms to screen for genes that, upon activation, confer resistance to a BRAF inhibitor. The top hits included genes previously shown to be able to confer resistance, and novel candidates were validated using individual sgRNA and complementary DNA overexpression. A gene expression signature based on the top screening hits correlated with markers of BRAF inhibitor resistance in cell lines and patient-derived samples. These results collectively demonstrate the potential of Cas9-based activators as a powerful genetic perturbation technology.
MIT Department
Harvard University--MIT Division of Health Sciences and Technology
Massachusetts Institute of Technology. Department of Biological Engineering
Massachusetts Institute of Technology. Department of Brain and Cognitive Sciences
McGovern Institute for Brain Research at MIT
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DOI of Published Version
https://doi.org/10.1038/nature14136
