High-resolution enzymatic mapping of genomic 5-hydroxymethylcytosine in mouse embryonic stem cells

Cell Rep. 2013 Feb 21;3(2):567-76. doi: 10.1016/j.celrep.2013.01.001. Epub 2013 Jan 24.


We describe the use of a unique DNA-modification-dependent restriction endonuclease AbaSI coupled with sequencing (Aba-seq) to map high-resolution hydroxymethylome of mouse E14 embryonic stem cells. The specificity of AbaSI enables sensitive detection of 5-hydroxymethylcytosine (5hmC) at low-occupancy regions. Bioinformatic analysis suggests 5hmCs in genic regions closely follow the 5mC distribution. 5hmC is generally depleted in CpG islands and only enriched in a small set of repetitive elements. A regularly spaced and oscillating 5hmC pattern was observed at the binding sites of CTCF. 5hmC is enriched at the poised enhancers with the monomethylated histone H3 lysine 4 (H3K4me1) marks, but not at the active enhancers with the acetylated histone H3 lysine 27 (H3K27Ac) marks. Non-CG hydroxymethylation appears to be prevalent in the mitochondrial genome. We propose that some amounts of transiently stable 5hmCs may indicate a poised epigenetic state or demethylation intermediate, whereas others may suggest a locally accessible chromosomal environment for the TET enzymatic apparatus.

Publication types

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

MeSH terms

  • 5-Methylcytosine / analogs & derivatives
  • Animals
  • Cell Line
  • Chromosome Mapping*
  • Computational Biology
  • CpG Islands
  • Cytosine / analogs & derivatives*
  • Cytosine / analysis
  • DNA Methylation
  • DNA Restriction Enzymes / metabolism*
  • Embryonic Stem Cells / cytology
  • Embryonic Stem Cells / metabolism*
  • Genomics
  • Histones / metabolism
  • Hydroxylation
  • Mice
  • Sequence Analysis, DNA*


  • Histones
  • 5-hydroxymethylcytosine
  • 5-Methylcytosine
  • Cytosine
  • DNA Restriction Enzymes

Associated data

  • GEO/GSE42898