What was once surprising is now established. DNA methylation is not static after imprinting. All genes are not silenced “forever” through DNA methylation. But how does this dynamic and reversible mechanism work? As the hypothesis goes, 5hmC is a step in the de-methylation process initiated by Tet dioxygenases. The news today is that 5hmC is more than just a quick step, it has function.
In Dynamic hydroxymethylation of deoxyribonucleic acid marks differentiation-associated enhancers. (2012) Nucleic Acids Research, 1-11. A.A. Serandour & S. Avner et al. show that conversion of 5mC to 5hmC activates enhancers – even pointing to it as an early step in the enhancer activation process. Can we all say functional signalling mark?
Some of you might be saying what are “enhancers” anyway? Enhancers are non-coding sequences which regulate long-distance gene expression. Their own activity is regulated by transcription factors. Enhancer sequences often vary significantly among different cell types. Although their sequences are not highly conserved, they can be ID’d through changes in the proteins that support DNA; histone marks, histone variant deposition and nucleosome stability.
For this study, samples of differentiating adipocyte cells and differentiating neural cells were run through a range of analyses, identifying new enhancers. hMeDIP-seq was used to select and analyze 5hmC genomic DNA. ChiP-seq was used to select for and analyze interactions among specific histone modifications, Tet1, and Meis1(a transcription factor). The FAIRE assay was used to demonstrate chromatin opening at enhancers. Transcriptome arrays were used to measure expression of differentiation genes. Newly ID’d enhancers were cloned into a luciferase reporter systems to establish their activities in vivo differentiating cells.
Data results were “heat maps of the relationships” at differentiation in the 2 cell line samples. I have to admit I’m terribly inadequate to explain the bioinformatics applied – see the paper for details. Simply put; When 5hmC is UP, hallmarks of ehancer activation H3K4me2 & H3K27ac are UP, Chromatin density is DOWN and differentiation associated gene expression levels are UP
Looks like 5hmC is an important signalling mark for initiating and maintaining cellular differentiation status, partially through regulation of enhancer activity. It makes sense since adult tissues and cells contain more 5hmC than both stem cells or most tumor tissues by immunolocalization. Cell specific hydroxymethylated enhancers are an influential layer of epigenetic instruction on DNA, attracting differentiation transcription factors and demethylating machinery like a magnet.
Sérandour AA, Avner S, Oger F, Bizot M, Percevault F, Lucchetti-Miganeh C, Palierne G, Gheeraert C, Barloy-Hubler F, Péron CL, Madigou T, Durand E, Froguel P, Staels B, Lefebvre P, Métivier R, Eeckhoute J, & Salbert G (2012). Dynamic hydroxymethylation of deoxyribonucleic acid marks differentiation-associated enhancers. Nucleic acids research PMID: 22730288