A set of genes critical to development is epigenetically poised in mouse germ cells from fetal stages through completion of meiosis

Abstract

In multicellular organisms, germ cells carry the hereditary material from one generation to the next. Developing germ cells are unipotent gamete precursors, and mature gametes are highly differentiated, specialized cells. However, upon gamete union at fertilization, their genomes drive a totipotent program, giving rise to a complete embryo as well as extraembryonic tissues. The biochemical basis for the ability to transition from differentiated cell to totipotent zygote is unknown. Here we report that a set of developmentally critical genes is maintained in an epigenetically poised (bivalent) state from embryonic stages through the end of meiosis. We performed ChIP-seq and RNA-seq analysis on flow-sorted male and female germ cells during embryogenesis at three time points surrounding sexual differentiation and female meiotic initiation, and then extended our analysis to meiotic and postmeiotic male germ cells. We identified a set of genes that is highly enriched for regulators of differentiation and retains a poised state (high H3K4me3, high H3K27me3, and lack of expression) across sexes and across developmental stages, including in haploid postmeiotic cells. The existence of such a state in embryonic stem cells has been well described. We now demonstrate that a subset of genes is maintained in a poised state in the germ line from the initiation of sexual differentiation during fetal development and into postmeiotic stages. We propose that the epigenetically poised condition of these developmental genes is a fundamental property of the mammalian germ-line nucleus, allowing differentiated gametes to unleash a totipotent program following fertilization.

Keywords: chromatin; epigenetics; spermatogenesis; totipotency.

Fig. 1.

ChIP-seq and RNA-seq data reflect the known biology of mouse fetal germ cells. (A) Sample gene tracks showing biological replicates of female E13.5 H3K4me3 and H3K27me3 data. (B) Correlated changes between H3K4me3 ChIP signal and gene expression for three core pluripotency regulators (Oct4, Sox2, and Nanog) and three genes associated with meiotic initiation (Stra8, Rec8, and Sycp3).

Fig. 2.

Clustering of H3K4me3 and H3K27me3 profiles with associated gene expression and functional enrichment. (A) Clustering of genes by similarity of H3K4me3 and H3K27me3 profiles 2 kb upstream and downstream of the transcriptional start site. (B) RNA expression levels for genes in each of four clusters. Each gray point represents a single gene; black dots, median; black lines, interquartile range. ***P < 10⁻¹⁵ (two-sample t test). (C) Top three enriched GO categories for each cluster.

Fig. 3.

A subset of poised genes is retained from E12.5 through E14.5. (A) Number of genes identified as “poised” at each fetal time point in male and female (gray bars), shared between time points in male and female (blue and red strips), and shared between time points and sexes (purple strip). (B) Distribution of poised genes identified in A among the four clusters identified in Fig. 2.

Fig. 4.

Promoters that are poised during fetal development remain poised during and after meiosis. (A) Schematic of meiosis in male germ cells. (B) Number of germ-line–poised genes retained across fetal time points in male only (blue) or in both male and female (purple), compared with genes identified as poised in both pachytene spermatocytes and round spermatids (“male meiotic,” green). (C) Sample gene tracks for three genes maintained in a poised state across germ cell development.

Fig. 5.

Germ-line–poised genes as a foundation for totipotency in the germ line. Critical developmental genes are continuously maintained in a poised state (purple and orange circles), with transcription repressed. Loss of either H3K4me3 (resolution toward the active state, with associated increases in gene expression) or H3K27me3 (resolution toward the repressed state) at specific promoters results in terminal differentiation. Those cells that maintain a complete set of poised developmental regulators throughout development constitute the germ cells.