EHMT2 directs DNA methylation for efficient gene silencing in mouse embryos.

The extent to which histone modifying enzymes contribute to DNA methylation in mammals remains unclear. Previous studies suggested a link between the lysine methyltransferase EHMT2 (also known as G9A and KMT1C) and DNA methylation in the mouse. Here, we used a model of knockout mice to explore the role of EHMT2 in DNA methylation during mouse embryogenesis. The Ehmt2 gene is expressed in epiblast cells but is dispensable for global DNA methylation in embryogenesis. In contrast, EHMT2 regulates DNA methylation at specific sequences that include CpG-rich promoters of germline-specific genes. These loci are bound by EHMT2 in embryonic cells, are marked by H3K9 dimethylation, and have strongly reduced DNA methylation in Ehmt2(-/-) embryos. EHMT2 also plays a role in the maintenance of germline-derived DNA methylation at one imprinted locus, the Slc38a4 gene. Finally, we show that DNA methylation is instrumental for EHMT2-mediated gene silencing in embryogenesis. Our findings identify EHMT2 as a critical factor that facilitates repressive DNA methylation at specific genomic loci during mammalian development.

E2F6 initiates stable epigenetic silencing of germline genes during embryonic development.

In mouse development, long-term silencing by CpG island DNA methylation is specifically targeted to germline genes; however, the molecular mechanisms of this specificity remain unclear. Here, we demonstrate that the transcription factor E2F6, a member of the polycomb repressive complex 1.6 (PRC1.6), is critical to target and initiate epigenetic silencing at germline genes in early embryogenesis. Genome-wide, E2F6 binds preferentially to CpG islands in embryonic cells. E2F6 cooperates with MGA to silence a subgroup of germline genes in mouse embryonic stem cells and in embryos, a function that critically depends on the E2F6 marked box domain. Inactivation of E2f6 leads to a failure to deposit CpG island DNA methylation at these genes during implantation. Furthermore, E2F6 is required to initiate epigenetic silencing in early embryonic cells but becomes dispensable for the maintenance in differentiated cells. Our findings elucidate the mechanisms of epigenetic targeting of germline genes and provide a paradigm for how transient repression signals by DNA-binding factors in early embryonic cells are translated into long-term epigenetic silencing during mouse development.

Genome-wide analysis in the mouse embryo reveals the importance of DNA methylation for transcription integrity.

Mouse embryos acquire global DNA methylation of their genome during implantation. However the exact roles of DNA methyltransferases (DNMTs) in embryos have not been studied comprehensively. Here we systematically analyze the consequences of genetic inactivation of Dnmt1, Dnmt3a and Dnmt3b on the methylome and transcriptome of mouse embryos. We find a strict division of function between DNMT1, responsible for maintenance methylation, and DNMT3A/B, solely responsible for methylation acquisition in development. By analyzing severely hypomethylated embryos, we uncover multiple functions of DNA methylation that is used as a mechanism of repression for a panel of genes including not only imprinted and germline genes, but also lineage-committed genes and 2-cell genes. DNA methylation also suppresses multiple retrotransposons and illegitimate transcripts from cryptic promoters in transposons and gene bodies. Our work provides a thorough analysis of the roles of DNA methyltransferases and the importance of DNA methylation for transcriptome integrity in mammalian embryos.

Ontogeny of CpG island methylation and specificity of DNMT3 methyltransferases during embryonic development in the mouse.

BACKGROUND: In the mouse, the patterns of DNA methylation are established during early embryonic development in the epiblast. We quantified the targets and kinetics of DNA methylation acquisition in epiblast cells, and determined the contribution of the de novo methyltransferases DNMT3A and DNMT3B to this process. RESULTS: We generated single-base maps of DNA methylation from the blastocyst to post-implantation stages and in embryos lacking DNMT3A or DNMT3B activity. DNA methylation is established within two days of implantation between embryonic days 4.5 and 6.5. The kinetics of de novo methylation are uniform throughout the genome, suggesting a random mechanism of deposition. In contrast, many CpG islands acquire methylation slowly in late epiblast cells. Five percent of CpG islands gain methylation and are found in the promoters of germline genes and in exons of important developmental genes. The onset of global methylation correlates with the upregulation of Dnmt3a/b genes in the early epiblast. DNMT3A and DNMT3B act redundantly to methylate the bulk genome and repetitive elements, whereas DNMT3B has a prominent role in the methylation of CpG islands on autosomes and the X chromosome. Reduced CpG island methylation in Dnmt3b-deficient embryos correlates with gene reactivation in promoters but reduced transcript abundance in gene bodies. Finally, DNMT3B establishes secondary methylation marks at imprinted loci, which distinguishes bona fide germline from somatic methylation imprints. CONCLUSIONS: We reveal that the DNMT3 de novo methyltransferases play both redundant and specific functions in the establishment of DNA methylation in the mouse embryo.

Mechanisms of DNA methylation and demethylation in mammals.

Cytosine methylation is an epigenetically propagated DNA modification that can modify how the DNA molecule is recognized and expressed. DNA methylation undergoes extensive reprogramming during mammalian embryogenesis and is directly linked to the regulation of pluripotency and cellular identity. Studying its regulation is also important for a better understanding of the many diseases that show epigenetic deregulations, in particular, cancer. In the recent years, a lot of progress has been made to characterize the profiles of DNA methylation at the genome level, which revealed that patterns of DNA methylation are highly dynamic between cell types. Here, we discuss the importance of DNA methylation for genome regulation and the mechanisms that remodel the DNA methylome during mammalian development, in particular the involvement of the rediscovered modified base 5-hydroxymethylcytosine.