DNA methylation is of paramount importance for mammalian embryonic development. DNA methylation has numerous functions: it is implicated in the repression of transposons and genes, but is also associated with actively transcribed gene bodies and, in some cases, with gene activation per se. In recent years, sensitive technologies have been developed that allow the interrogation of DNA methylation patterns from a small number of cells. The use of these technologies has greatly improved our knowledge of DNA methylation dynamics and heterogeneity in embryos and in specific tissues. Combined with genetic analyses, it is increasingly apparent that regulation of DNA methylation erasure and (re-)establishment varies considerably between different developmental stages. In this Review, we discuss the mechanisms and functions of DNA methylation and demethylation in both mice and humans at CpG-rich promoters, gene bodies and transposable elements. We highlight the dynamic erasure and re-establishment of DNA methylation in embryonic, germline and somatic cell development. Finally, we provide insights into DNA methylation gained from studying genetic diseases. DNA methylation is essential for mammalian embryogenesis owing to its repression of transposons and genes, but it is also associated with gene activation. The recent use of sensitive technologies has revealed that DNA methylation dynamics vary considerably between embryonic, germline and somatic cell development, with implications for genetic diseases and cancer.

The diverse roles of DNA methylation in mammalian development and disease

A key mode of regulating DNA methylation is through active demethylation driven by TET-mediated oxidation of 5-methylcytosine (5mC). This Review discusses our latest understanding of the mechanisms and regulation of active DNA demethylation, and the roles of active demethylation (and the oxidized 5mC intermediates) in gene regulation, genome stability, development and disease.

TET-mediated active DNA demethylation: mechanism, function and beyond

The pattern of DNA methylation at cytosine bases in the genome is tightly linked to gene expression, and DNA methylation abnormalities are often observed in diseases. The ten eleven translocation (TET) enzymes oxidize 5-methylcytosines (5mCs) and promote locus-specific reversal of DNA methylation. TET genes, and especially TET2, are frequently mutated in various cancers, but how the TET proteins contribute to prevent the onset and maintenance of these malignancies is largely unknown. Here, we highlight recent advances in understanding the physiological function of the TET proteins and their role in regulating DNA methylation and transcription. In addition, we discuss some of the key outstanding questions in the field.

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Role of TET enzymes in DNA methylation, development, and cancer

In mammals, extensive chromatin reorganization is essential for reprogramming terminally committed gametes to a totipotent state during preimplantation development. However, the global chromatin landscape and its dynamics in this period remain unexplored. Here we report a genome-wide map of accessible chromatin in mouse preimplantation embryos using an improved assay for transposase-accessible chromatin with high throughput sequencing (ATAC-seq) approach with CRISPR/Cas9-assisted mitochondrial DNA depletion. We show that despite extensive parental asymmetry in DNA methylomes, the chromatin accessibility between the parental genomes is globally comparable after major zygotic genome activation (ZGA). Accessible chromatin in early embryos is widely shaped by transposable elements and overlaps extensively with putative cis-regulatory sequences. Unexpectedly, accessible chromatin is also found near the transcription end sites of active genes. By integrating the maps of cis-regulatory elements and single-cell transcriptomes, we construct the regulatory network of early development, which helps to identify the key modulators for lineage specification. Finally, we find that the activities of cis-regulatory elements and their associated open chromatin diminished before major ZGA. Surprisingly, we observed many loci showing non-canonical, large open chromatin domains over the entire transcribed units in minor ZGA, supporting the presence of an unusually permissive chromatin state. Together, these data reveal a unique spatiotemporal chromatin configuration that accompanies early mammalian development.

The landscape of accessible chromatin in mammalian preimplantation embryos

Three papers in this issue of Nature use highly sensitive ChIP–seq assays to describe the dynamic patterns of histone modifications during early mouse embryogenesis, showing that oocytes have a distinctive epigenome and providing insights into how the maternal gene expression program transitions to the zygotic program. Genomic analysis of chromatin states in early embryos has been technically difficult, owing to the limited number of cells available for analysis. Three papers in this issue of Nature use highly sensitive ChIP–seq assays to describe the dynamic patterns of histone modifications during early mouse embryogenesis. Arne Klungland and colleagues find that the oocyte genome is associated with broad non-canonical domains of histone H3K4me3 which seem to function in preventing deposition of DNA methylation. Wei Xie and colleagues find that the oocyte genome is associated with broad non-canonical domains of histone H3K4me3 which overlap with domains of low DNA methylation and seem to contribute to gene silencing. Shaorong Gao and colleagues map histone H3K4me3 and H3K27me3 modifications in pre-implantation embryos and focus on the re-establishment of histone modifications during zygotic genome activation. They find that the breadth of H3K4me3 domains is highly dynamic and that H3K4me3 re-establishes rapidly on promoter regions whereas H3K27me3 is mostly absent from these regions. Taken together—and with previously published work—these studies show that the oocyte has a distinctive epigenome and provide insights into how the maternal gene expression program transitions to the zygotic program. Histone modifications have critical roles in regulating the expression of developmental genes during embryo development in mammals1,2. However, genome-wide analyses of histone modifications in pre-implantation embryos have been impeded by the scarcity of the required materials. Here, by using a small-scale chromatin immunoprecipitation followed by sequencing (ChIP–seq) method3, we map the genome-wide profiles of histone H3 lysine 4 trimethylation (H3K4me3) and histone H3 lysine 27 trimethylation (H3K27me3), which are associated with gene activation and repression4,5, respectively, in mouse pre-implantation embryos. We find that the re-establishment of H3K4me3, especially on promoter regions, occurs much more rapidly than that of H3K27me3 following fertilization, which is consistent with the major wave of zygotic genome activation at the two-cell stage. Furthermore, H3K4me3 and H3K27me3 possess distinct features of sequence preference and dynamics in pre-implantation embryos. Although H3K4me3 modifications occur consistently at transcription start sites, the breadth of the H3K4me3 domain is a highly dynamic feature. Notably, the broad H3K4me3 domain (wider than 5 kb) is associated with higher transcription activity and cell identity not only in pre-implantation development but also in the process of deriving embryonic stem cells from the inner cell mass and trophoblast stem cells from the trophectoderm. Compared to embryonic stem cells, we found that the bivalency (that is, co-occurrence of H3K4me3 and H3K27me3) in early embryos is relatively infrequent and unstable. Taken together, our results provide a genome-wide map of H3K4me3 and H3K27me3 modifications in pre-implantation embryos, facilitating further exploration of the mechanism for epigenetic regulation in early embryos.

Neel Aluru

Papers

Programming and Inheritance of Parental DNA Methylomes in Mammals