Repression of germline genes by PRC1.6 and SETDB1 in the early embryo precedes DNA methylation-mediated silencing

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.

Nucleome programming is required for the foundation of totipotency in mammalian germline development

“…These genes, which included many subject to insulation erasure upon spermatogonia development (Fig 3I and J), are referred to as “germline genes” (Borgel et al , 2010), and are known to be repressed by DNA methylation in somatic cells and by H3K27me3 and H3K9me2 in mPGCLCs (Borgel et al , 2010; Kurimoto et al , 2015). Furthermore, a recent report has shown that the germline genes were repressed in EpiLCs with H3K9me3 imposed by Setdb1 (Mochizuki et al , 2021). In good agreement, the transcriptional start sites (TSSs) of germline genes repressed by Setdb1 up‐regulated H3K9me3 in EpiLCs and, more prominently, in d2 mPGCLCs, and lost it in d4c7 mPGCLCs (Fig 5K and L).…”

Rif1 interacts with non-canonical polycomb repressive complex PRC1.6 to regulate mouse embryonic stem cells fate potential

“…The functional differences of the PRC1.6 complex resulted from these distinct recruitment mechanisms are of interest to be elaborated in the future. The PRC1.6 complex inhibits premature differentiation of mESCs by repressing germ cell-related genes (Dahlet et al 2021 ; Endoh et al 2017 ; Liu et al 2020 ; Maeda et al 2013 ; Mochizuki et al 2021 ; Suzuki et al 2016 ; Uranishi et al 2021 ), and loss of Max results in meiotic entry even in mESCs (Suzuki et al 2016 ). Similar to that of Pcgf6, we found downregulation of Rif1 also caused mis-regulation of genes in meiosis-related pathways (Fig.…”

“…Each contains a distinct Pcgf subunit (Gao et al 2012 ). The PRC1.6 complex, consisting of multiple subunits including Pcgf6, RNF2, RYBP, L3mbtl2, Mga, Max, and E2F6, is known to inhibit meiotic entry of embryonic cells by targeting meiosis and germline genes (Dahlet et al 2021 ; Endoh et al 2017 ; Liu et al 2020 ; Maeda et al 2013 ; Mochizuki et al 2021 ; Suzuki et al 2016 ; Uranishi et al 2021 ). Depletion of PRC1.6 subunits such as Pcgf6 or L3mbtl2 results in multiple defects in embryonic development, including failure of gastrulation, abnormal axis development, and embryonic lethality (Endoh et al 2017 ; Liu et al 2020 ; Qin et al 2012 ).…”

Intragenic CpG Islands and Their Impact on Gene Regulation

“…A more stable form of repression at CGIs is through DNA methylation. In somatic tissues, DNA methylation represses promoter CGIs at the inactivated X chromosome (Augui et al, 2011;Galupa and Heard, 2018), germline genes (Velasco et al, 2010;Dahlet et al, 2021;Mochizuki et al, 2021), imprinted genes (Barlow and Bartolomei, 2014), and some lineage-committed genes (Dahlet et al, 2020). Whilst H3K27me3 and DNA methylation are both repressive, they are typically mutually exclusive at CGIs (Brinkman et al, 2012;Statham et al, 2012).…”