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Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complex
  • Published Date:
    Dec 10 2014
  • Source:
    Nature. 517(7536):583-588.
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Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complex
Details:
  • Description:
    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.

  • Document Type:
  • Collection(s):
  • Funding:
    DP1 MH100706/MH/NIMH NIH HHS/United States
    DP1-MH100706/DP/NCCDPHP CDC HHS/United States
    R01 NS062849/NS/NINDS NIH HHS/United States
    R01 NS073124/NS/NINDS NIH HHS/United States
    R01-NS07312401/NS/NINDS NIH HHS/United States
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