DNA methylation programming and reprogramming in primate embryonic stem cells

Abstract

DNA methylation is an important epigenetic mechanism, affecting normal development and playing a key role in reprogramming epigenomes during stem cell derivation. Here we report on DNA methylation patterns in native monkey embryonic stem cells (ESCs), fibroblasts, and ESCs generated through somatic cell nuclear transfer (SCNT), identifying and comparing epigenome programming and reprogramming. We characterize hundreds of regions that are hyper-or hypomethylated in fibroblasts compared to native ESCs and show that these are conserved in human cells and tissues. Remarkably, the vast majority of these regions are reprogrammed in SCNT ESCs, leading to almost perfect correlation between the epigenomic profiles of the native and reprogrammed lines. At least 58% of these changes are correlated in cis to transcription changes, Polycomb Repressive Complex-2 occupancy, or binding by the CTCF insulator. We also show that while epigenomic reprogramming is extensive and globally accurate, the efficiency of adding and stripping DNA methylation during reprogramming is regionally variable. In several cases, this variability results in regions that remain methylated in a fibroblast-like pattern even after reprogramming.[Supplemental material is available online at http://www.genome.org. DNA methylation profiles from this study have been submitted to the Gene Expression Omnibus (http://www.ncbi.nlm.nih.gov/geo) under accession no. GSE17981.]DNA methylation is considered a key factor in the formation of cellular memory and identity, but owing to experimental and conceptual limitations, we still do not truly understand how the cell writes and erases DNA methylation marks in the course of normal cellular differentiation, and how these marks revert to their original embryonic stem cell (ESC)-like form following somatic cell nuclear transfer or induced pluripotent stem cell (iPS) reprogramming (Reik 2007). Progress in the field was hampered for years by lack of quality methods for high-throughput DNA methylation profiling, but recently several effective assays for profiling DNA methylation in large fractions of the mammalian genome were developed and applied successfully (Weber et al. 2005;Keshet et al. 2006;Rollins et al. 2006;Cokus et al. 2008;Irizarry et al. 2008;Meissner et al. 2008). Another major source of confusion and difficulty in understanding the role of mammalian DNA methylation is the nonuniform CpG content of the genome, which led most of the experimental attention toward regions with high CpG content (CpG islands). Recent evidence suggests that classical CpG islands of high CpG content are almost never methylated under normal conditions, yet much dynamic DNA methylation (manifested as differentially methylated regions, DMRs) can be found in regions with intermediate CpG content, some of which are classically defined as CpG islands and some of which are not (Irizarry et al. 2009;Straussman et al. 2009). Adding to these difficulties, multiple studies have shown that DNA methylation is stably acquired in culture, forming s...

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“…Contrasting LMRs with maps of histone modifications, DNase I hypersensitivity and several DNA-binding factors revealed that LMRs represent distal regulatory regions, such as enhancers. This is in agreement with several single-gene studies of methylation dynamics at localized distal regulatory regions 56, 57. Most recently, Hon et al 58 identified 302,864 tissue-specific differentially methylated regions (tsDMRs) by profiling the methylomes of a diverse panel of 17 normal adult mouse tissues.…”

Epigenetic polymorphism and the stochastic formation of differentially methylated regions in normal and cancerous tissues

“…Previous studies in ESCs, differentiation models, cancer cell lines and tumors have used popu lationaveraged measurements to show that H3K4me3, H3K27me3 and CTCF binding are all spatially and temporally anticorrelated with DNA methylation in regions with high CpG content 26,[44][45][46][47][48] . In the in vitro system studied here, we recapitulate these reciprocal relation ships.…”

Systems biology discoveries using non-human primate pluripotent stem and germ cells: novel gene and genomic imprinting interactions as well as unique expression patterns

“…One such epigenetic mechanism is DNA methylation, which is considered a key factor in the formation of cellular memory and identity [129]. A comprehensive review summarized the key features of the regulatory mechanisms that control the transcriptional regulatory features in hESCs [130,131], which complemented their work with ChIP-chip in mESCs [132].…”

“…In several cases, this variability results in regions that remain methylated in a fibroblast-like pattern even after reprogramming [129]. …”