Highly specific epigenome editing by CRISPR-Cas9 repressors for silencing of distal regulatory elements

Nat Methods. 2015 Dec;12(12):1143-9. doi: 10.1038/nmeth.3630. Epub 2015 Oct 26.

Abstract

Epigenome editing with the CRISPR (clustered, regularly interspaced, short palindromic repeats)-Cas9 platform is a promising technology for modulating gene expression to direct cell phenotype and to dissect the causal epigenetic mechanisms of gene regulation. Fusions of nuclease-inactive dCas9 to the Krüppel-associated box (KRAB) repressor (dCas9-KRAB) can silence target gene expression, but the genome-wide specificity and the extent of heterochromatin formation catalyzed by dCas9-KRAB are not known. We targeted dCas9-KRAB to the HS2 enhancer, a distal regulatory element that orchestrates the expression of multiple globin genes, and observed highly specific induction of H3K9 trimethylation (H3K9me3) at the enhancer and decreased chromatin accessibility of both the enhancer and its promoter targets. Targeted epigenetic modification of HS2 silenced the expression of multiple globin genes, with minimal off-target changes in global gene expression. These results demonstrate that repression mediated by dCas9-KRAB is sufficiently specific to disrupt the activity of individual enhancers via local modification of the epigenome.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • CRISPR-Associated Proteins / genetics*
  • CRISPR-Cas Systems / genetics*
  • Clustered Regularly Interspaced Short Palindromic Repeats / genetics*
  • Enhancer Elements, Genetic
  • Epigenesis, Genetic*
  • Epigenomics / methods*
  • Gene Expression Regulation, Viral
  • Globins / genetics
  • HEK293 Cells
  • Humans
  • K562 Cells
  • Lentivirus / genetics
  • RNA, Guide, CRISPR-Cas Systems / genetics
  • Regulatory Elements, Transcriptional / genetics*
  • Repressor Proteins / genetics
  • Viral Proteins / genetics

Substances

  • CRISPR-Associated Proteins
  • RNA, Guide, CRISPR-Cas Systems
  • Repressor Proteins
  • Viral Proteins
  • Globins