Abstract
Silencing of a subset of germline genes is dependent upon DNA methylation (DNAme) post-implantation. However, these genes are generally hypomethylated in the blastocyst, implicating alternative repressive pathways before implantation. Indeed, in embryonic stem cells (ESCs), an overlapping set of genes, including germline “genome-defence” (GGD) genes, are upregulated following deletion of the H3K9 methyltransferase SETDB1 or subunits of the non-canonical PRC1 complex PRC1.6. Here, we show that in pre-implantation embryos and naïve ESCs (nESCs), hypomethylated promoters of germline genes bound by the PRC1.6 DNA-binding subunits MGA/MAX/E2F6 are enriched for RING1B-dependent H2AK119ub1 and H3K9me3. Accordingly, repression of these genes in nESCs shows a greater dependence on PRC1.6 than DNAme. In contrast, GGD genes are hypermethylated in epiblast-like cells (EpiLCs) and their silencing is dependent upon SETDB1, PRC1.6/RING1B and DNAme, with H3K9me3 and DNAme establishment dependent upon MGA binding. Thus, GGD genes are initially repressed by PRC1.6, with DNAme subsequently engaged in post-implantation embryos.
Highlights
This epigenetic mark is highly dynamic during embryonic development, with a genome-wide wave of demethylation occurring after fertilization, mediated at least in part by dioxygenases of the ten-eleven translocation (TET) protein family[2]
The DNA methylation (DNAme) level of these MGA- and E2F6-bound genes is generally lower than that of the DMS germline genes that are not bound by these transcription factors (TFs), a trend observed in E8.5 embryos (Fig. 1b and Supplementary Fig. 1a)
By integrating numerous previously published datasets from preimplantation stages, we show that the promoter regions of DMS germline genes bound by MGA/MAX and E2F6 in embryonic stem cells (ESCs) show sequential accumulation of H2AK119ub[1], H3K9me[3], and H3K27me[3], followed by de novo DNAme in the epiblast (Fig. 6a)
Summary
This epigenetic mark is highly dynamic during embryonic development, with a genome-wide wave of demethylation occurring after fertilization, mediated at least in part by dioxygenases of the ten-eleven translocation (TET) protein family[2]. While DNAme is generally maintained at high levels in somatic lineages[5], another wave of demethylation occurs early in germ cell development in both males and females[6]. During this period, many genes, including those essential for meiosis and gametogenesis, are upregulated. 137 germline genes that acquire dense promoter DNAme in wild-type (WT) embryos by E6.5 are derepressed in Dnmt3a/3b double knock-out (DKO) embryos at E8.512. This list of DNAme-sensitive (DMS) germline genes includes many that encode proteins involved in the repression of transposable elements (TEs), such as piRNA biogenesis factors. While it has been speculated that DNMT3B recruitment through E2F6 transcriptional repressor mediates germline gene silencing in mouse somatic tissues[22], DNMT3A and DNMT3B act in a redundant manner to methylate the CGI promoters of such genes in peri-implantation embryos[4]
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