title: CRISPR-mediated modular RNA-guided regulation of transcription in eukaryotes.
abstract: The genetic interrogation and reprogramming of cells requires methods for robust and precise targeting of genes for expression or repression. The CRISPR-associated catalytically inactive dCas9 protein offers a general platform for RNA-guided DNA targeting. Here, we show that fusion of dCas9 to effector domains with distinct regulatory functions enables stable and efficient transcriptional repression or activation in human and yeast cells, with the site of delivery determined solely by a coexpressed short guide (sg)RNA. Coupling of dCas9 to a transcriptional repressor domain can robustly silence expression of multiple endogenous genes. RNA-seq analysis indicates that CRISPR interference (CRISPRi)-mediated transcriptional repression is highly specific. Our results establish that the CRISPR system can be used as a modular and flexible DNA-binding platform for the recruitment of proteins to a target DNA sequence, revealing the potential of CRISPRi as a general tool for the precise regulation of gene expression in eukaryotic cells.
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: Luke A
authors.initials: LA
authors.lastname: Gilbert
authors.forename: Matthew H
authors.initials: MH
authors.lastname: Larson
authors.forename: Leonardo
authors.initials: L
authors.lastname: Morsut
authors.forename: Zairan
authors.initials: Z
authors.lastname: Liu
authors.forename: Gloria A
authors.initials: GA
authors.lastname: Brar
authors.forename: Sandra E
authors.initials: SE
authors.lastname: Torres
authors.forename: Noam
authors.initials: N
authors.lastname: Stern-Ginossar
authors.forename: Onn
authors.initials: O
authors.lastname: Brandman
authors.forename: Evan H
authors.initials: EH
authors.lastname: Whitehead
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Doudna
authors.forename: Wendell A
authors.initials: WA
authors.lastname: Lim
authors.forename: Jonathan S
authors.initials: JS
authors.lastname: Weissman
authors.forename: Lei S
authors.initials: LS
authors.lastname: Qi
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics*; D018390:Gene Targeting / Q000379:methods*; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013297:Streptococcus pyogenes; D058727:RNA, Small Untranslated
chemical_list: D001426:Bacterial Proteins; D058727:RNA, Small Untranslated