Identification of in vivo DNA targets of chromatin proteins using tethered dam methyltransferase.
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.
van Steensel
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B
Fred Hutchinson Cancer Research Center and Howard Hughes Medical Institute, Seattle, WA 98109, USA. bsteense@fhcrc.org
Henikoff
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S
D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019175:DNA Methylation; D004331:Drosophila melanogaster; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins; D005821:Genetic Techniques; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D016366:Open Reading Frames; D016133:Polymerase Chain Reaction / Q000379:methods; D011485:Protein Binding; D015183:Restriction Mapping; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000235:genetics / Q000378:metabolism*; D014162:Transfection / Q000379:methods
D002843:Chromatin; D029968:Escherichia coli Proteins; D009687:Nuclear Proteins; D004247:DNA; C058693:Dam methyltransferase; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific); C499294:dam protein, E coli