title: Tissue-specific distribution and dynamic changes of 5-hydroxymethylcytosine in mammalian genomes.
abstract: Cytosine residues in the vertebrate genome are enzymatically modified to 5-methylcytosine, which participates in transcriptional repression of genes during development and disease progression. 5-Methylcytosine can be further enzymatically modified to 5-hydroxymethylcytosine by the TET family of methylcytosine dioxygenases. Analysis of 5-methylcytosine and 5-hydroxymethylcytosine is confounded, as these modifications are indistinguishable by traditional sequencing methods even when supplemented by bisulfite conversion. Here we demonstrate a simple enzymatic approach that involves cloning, identification, and quantification of 5-hydroxymethylcytosine in various CCGG loci within murine and human genomes. 5-Hydroxymethylcytosine was prevalent in human and murine brain and heart genomic DNAs at several regions. The cultured cell lines NIH3T3 and HeLa both displayed very low or undetectable amounts of 5-hydroxymethylcytosine at the examined loci. Interestingly, 5-hydroxymethylcytosine levels in mouse embryonic stem cell DNA first increased then slowly decreased upon differentiation to embryoid bodies, whereas 5-methylcytosine levels increased gradually over time. Finally, using a quantitative PCR approach, we established that a portion of VANGL1 and EGFR gene body methylation in human tissue DNA samples is indeed hydroxymethylation.
authors: New England Biolabs, Inc., Ipswich, Massachusetts 01938, USA.
authors: Shannon Morey
authors: SM
authors: Kinney
authors: Hang Gyeong
authors: HG
authors: Chin
authors: Romualdas
authors: R
authors: Vaisvila
authors: Jurate
authors: J
authors: Bitinaite
authors: Yu
authors: Y
authors: Zheng
authors: Pierre-Olivier
authors: PO
authors: Estève
authors: Suhua
authors: S
authors: Feng
authors: Hume
authors: H
authors: Stroud
authors: Steven E
authors: SE
authors: Jacobsen
authors: Sriharsa
authors: S
authors: Pradhan
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