Developmentally programmed 3' CpG island methylation confers tissue- and cell-type-specific transcriptional activation

Mol Cell Biol. 2013 May;33(9):1845-58. doi: 10.1128/MCB.01124-12. Epub 2013 Mar 4.


During development, a small but significant number of CpG islands (CGIs) become methylated. The timing of developmentally programmed CGI methylation and associated mechanisms of transcriptional regulation during cellular differentiation, however, remain poorly characterized. Here, we used genome-wide DNA methylation microarrays to identify epigenetic changes during human embryonic stem cell (hESC) differentiation. We discovered a group of CGIs associated with developmental genes that gain methylation after hESCs differentiate. Conversely, erasure of methylation was observed at the identified CGIs during subsequent reprogramming to induced pluripotent stem cells (iPSCs), further supporting a functional role for the CGI methylation. Both global gene expression profiling and quantitative reverse transcription-PCR (RT-PCR) validation indicated opposing effects of CGI methylation in transcriptional regulation during differentiation, with promoter CGI methylation repressing and 3' CGI methylation activating transcription. By studying diverse human tissues and mouse models, we further confirmed that developmentally programmed 3' CGI methylation confers tissue- and cell-type-specific gene activation in vivo. Importantly, luciferase reporter assays provided evidence that 3' CGI methylation regulates transcriptional activation via a CTCF-dependent enhancer-blocking mechanism. These findings expand the classic view of mammalian CGI methylation as a mechanism for transcriptional silencing and indicate a functional role for 3' CGI methylation in developmental gene regulation.

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
  • CCCTC-Binding Factor
  • Cell Differentiation
  • Cell Line
  • CpG Islands*
  • DNA / genetics
  • DNA Methylation*
  • Embryonic Stem Cells / cytology*
  • Embryonic Stem Cells / metabolism*
  • Epigenesis, Genetic
  • Gene Expression Regulation, Developmental*
  • Humans
  • Induced Pluripotent Stem Cells / cytology
  • Induced Pluripotent Stem Cells / metabolism
  • Methylation
  • Mice
  • Mice, Inbred C57BL
  • Oligonucleotide Array Sequence Analysis
  • Repressor Proteins / metabolism
  • Transcriptional Activation*


  • CCCTC-Binding Factor
  • CTCF protein, human
  • Ctcf protein, mouse
  • Repressor Proteins
  • DNA