Analysis of steric effects in DamID profiling of transcription factor target genes

Mirana Ramialison, Ashley J. Waardenberg, Nicole Schonrock, Tram Doan, Danielle de Jong, Romaric Bouveret, Richard P. Harvey

Research output: Contribution to journalArticleResearchpeer-review

3 Citations (Scopus)

Abstract

DNA adenine methyltransferase identification (DamID) is an enzymatic technology for detecting DNA regions targeted by chromatin-associated proteins. Proteins are fused to bacterial DNA adenine methyltransferase (Dam) and expressed in cultured cells or whole organisms. Here, we used DamID to detect DNA regions bound by the cardiac-restricted transcription factors (TFs) NKX2-5 and SRF, and ubiquitously-expressed co-factors ELK1 and ELK4. We compared targets bound by these TFs as N- and C-terminal fusions with Dam, for both wild type (WT) NKX2-5 and mutant proteins mimicking those found in congenital heart disease. Overall, DamID is highly robust: while the orientation of WT Dam fusions can affect the size of the target sets, their signatures remained largely reproducible. Furthermore, a severe NKX2-5 mutant lacking the homeodomain showed strong steric effects negatively impacting target discovery. The extent of steric effect is likely to be dependent on the protein in question and the orientation of Dam fusion.

Original languageEnglish
Pages (from-to)75-82
Number of pages8
JournalGenomics
Volume109
Issue number2
DOIs
Publication statusPublished - 1 Mar 2017

Keywords

  • Congenital heart disease
  • DNA adenine methyltransferase identification (DamID)
  • NKX2-5
  • Quality control
  • Steric effect

Cite this

Ramialison, M., Waardenberg, A. J., Schonrock, N., Doan, T., de Jong, D., Bouveret, R., & Harvey, R. P. (2017). Analysis of steric effects in DamID profiling of transcription factor target genes. Genomics, 109(2), 75-82. https://doi.org/10.1016/j.ygeno.2017.01.006