Genome-wide tracking of dCas9-methyltransferase footprints

Nat Commun. 2018 Feb 9;9(1):597. doi: 10.1038/s41467-017-02708-5.

Abstract

In normal mammalian development cytosine methylation is essential and is directed to specific regions of the genome. Despite notable advances through mapping its genome-wide distribution, studying the direct contribution of DNA methylation to gene and genome regulation has been limited by the lack of tools for its precise manipulation. Thus, combining the targeting capability of the CRISPR-Cas9 system with an epigenetic modifier has attracted interest in the scientific community. In contrast to profiling the genome-wide cleavage of a nuclease competent Cas9, tracing the global activity of a dead Cas9 (dCas9) methyltransferase fusion protein is challenging within a highly methylated genome. Here, we report the generation and use of an engineered, methylation depleted but maintenance competent mouse ES cell line and find surprisingly ubiquitous nuclear activity of dCas9-methyltransferases. Subsequent experiments in human somatic cells refine these observations and point to an important difference between genetic and epigenetic editing tools that require unique experimental considerations.

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

  • Animals
  • Bacterial Proteins
  • CRISPR-Associated Protein 9
  • Cell Line*
  • DNA (Cytosine-5-)-Methyltransferases / metabolism*
  • Embryonic Stem Cells / enzymology*
  • Endonucleases
  • Gene Editing
  • Humans
  • Mice

Substances

  • Bacterial Proteins
  • DNA (Cytosine-5-)-Methyltransferases
  • CRISPR-Associated Protein 9
  • Cas9 endonuclease Streptococcus pyogenes
  • Endonucleases