Identification of in Vivo DNA Targets of Chromatin Proteins Using Tethered Dam Methyltransferase

Nat Biotechnol. 2000 Apr;18(4):424-8. doi: 10.1038/74487.

Abstract

We have developed a novel technique, named DamID, for the identification of DNA loci that interact in vivo with specific nuclear proteins in eukaryotes. By tethering Escherichia coli DNA adenine methyltransferase (Dam) to a chromatin protein, Dam can be targeted in vivo to native binding sites of this protein, resulting in local DNA methylation. Sites of methylation can subsequently be mapped using methylation-specific restriction enzymes or antibodies. We demonstrate the successful application of DamID both in Drosophila cell cultures and in whole flies. When Dam is tethered to the DNA-binding domain of GAL4, targeted methylation is limited to a region of a few kilobases surrounding a GAL4 binding sequence. Using DamID, we identified a number of expected and unexpected target loci for Drosophila heterochromatin protein 1. DamID has potential for genome-wide mapping of in vivo targets of chromatin proteins in various eukaryotes.

MeSH terms

  • Animals
  • Cell Line
  • Chromatin / metabolism*
  • DNA / chemistry
  • DNA / genetics
  • DNA / metabolism*
  • DNA Methylation
  • Drosophila melanogaster
  • Escherichia coli / enzymology
  • Escherichia coli Proteins
  • Genetic Techniques
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism*
  • Open Reading Frames
  • Polymerase Chain Reaction / methods
  • Protein Binding
  • Restriction Mapping
  • Site-Specific DNA-Methyltransferase (Adenine-Specific) / genetics
  • Site-Specific DNA-Methyltransferase (Adenine-Specific) / metabolism*
  • Transfection / methods

Substances

  • Chromatin
  • Escherichia coli Proteins
  • Nuclear Proteins
  • DNA
  • Dam methyltransferase
  • Site-Specific DNA-Methyltransferase (Adenine-Specific)
  • dam protein, E coli