Tissue-specific demethylation in CpG-poor promoters during cellular differentiation

Abstract

Epigenetic regulation is essential in determining cellular phenotypes during differentiation. Although tissue-specific DNA methylation has been studied, the significance of methylation variance for tissue phenotypes remains unresolved, especially for CpG-poor promoters. Here, we comprehensively studied methylation levels of 27 578 CpG sites among 21 human normal tissues from 12 anatomically different regions using an epigenotyping beadarray system. Remarkable changes in tissue-specific DNA methylation were observed within CpG-poor promoters but not CpG-rich promoters. Of note, tissue-specific hypomethylation is accompanied by an increase in gene expression, which gives rise to specialized cellular functions. The hypomethylated regions were significantly enriched with recognition motifs for transcription factors that regulate cell-type-specific differentiation. To investigate the dynamics of hypomethylation events, we analyzed methylation levels of the entire APOA1 gene locus during in vitro differentiation of embryonic stem cells toward the hepatic lineage. A decrease in methylation was observed after day 13, coinciding with alpha-fetoprotein detection, in the vicinity of its transcription start sites (TSSs), and extends up to ∼200 bp region encompassing the TSS at day 21, equivalent to the hepatoblastic stage. This decrease is even more pronounced in the adult liver, where the entire APOA1 gene locus is hypomethylated. Furthermore, when we compared the methylation status of induced pluripotent stem (iPS) cells with their parental cell, IMR-90, we found that fibroblast-specific hypomethylation is restored to a fully methylated state in iPS cells after reprogramming. These results illuminate tissue-specific methylation dynamics in CpG-poor promoters and provide more comprehensive views on spatiotemporal gene regulation in terminal differentiation.

A DNA methylation fingerprint of 1628 human samples

“…Herein, using the first version of the DNA methylation bead microarray, which included 1505 CpG sites corresponding to 808 genes, we have studied the largest collection of human samples to date, 1628, that included 424 normal tissues, 1054 tumorigenic samples, and 150 non-cancerous disorders. Our data provide new clues about the DNA methylation profiles present in normal and diseaseassociated tissues and also expand and confirm previous reports in this area obtained using the same platform (Aranda et al 2009;Byun et al 2009;Christensen et al 2009;Javierre et al 2010) or a second DNA methylation bead microarray that includes 27,000 CpG sites (Rakyan et al 2010;Teschendorff et al 2010;Nagae et al 2011). In normal cells, the derived picture reinforces the role of methylation in non-CpG-island 59 ends to determine tissue-specific expression, the shift in the DNA methylation landscape from pluripotent to differentiated cells, and the existence of a DNA methylation drift associated with aging.…”

“…1A). The 511 CpG sites described correspond to 359 genes and, providing further validation to the data, 220 genes (61%; 220) and 137 (38%) were previously identified as genes with tissue-specific DNA methylation using the same 1505 CpG platform (Byun et al 2009) or a 27,000-CpG microarray (Nagae et al 2011), respectively. Illustrative examples of genes found in the three sets, and also confirmed by bisulfite genomic sequencing in another independent study (Eckhardt et al 2006), include TBX1 (T-box 1), OSM (oncostatin M), and GP1BB (glycoprotein Ib [platelet] beta polypeptide).…”

What Do You Mean, “Epigenetic”?

“…This is particularly true when the attempts to validate these hypotheses have, as of yet, proved inconclusive. What can it mean to say that DNA methylation is repressive when activation of a gene removes methylation (e.g., Bird 2002;Nagae et al 2011;Hackett et al 2012;Qian et al 2012;Gan et al 2013;Xie et al 2013;Bestor et al 2014)? The search for the mechanism of semiconservative histone modifications continues (Deal et al 2010;Xu et al 2010;Nakano et al 2011;Tran et al 2012;Whitehouse and Smith 2013) despite evidence that the modifications respond to expression state rather than control it (Kilpinen et al 2013;Ptashne 2014;Teves et al 2014).…”

Generation of induced pluripotent stem cells from primary chronic myelogenous leukemia patient samples

“…Methylation status was evaluated as previously reported. 28 Methylation status was analyzed using HumanMethylation27 BeadChip (Illumina). Genomic DNA for methylation profiling was quantified using the Quant-iT dsDNA BR assay kit (Invitrogen).…”