Maternal-to-zygotic transition (MZT) is essential for the formation of a new individual, but is still poorly understood despite recent progress in analysis of gene expression and DNA methylation in early embryogenesis1–9. Dynamic histone modifications may have important roles in MZT10–13, but direct measurements of chromatin states have been hindered by technical difficulties in profiling histone modifications from small quantities of cells. Recent improvements allow for 500 cell-equivalents of chromatin per reaction, but require 10,000 cells for initial steps14 or require a highly specialized microfluidics device that is not readily available15. We developed a micro-scale chromatin immunoprecipitation and sequencing (μChlP-seq) method, which we used to profile genome-wide histone H3 lysine methylation (H3K4me3) and acetylation (H3K27ac) in mouse immature and metaphase II oocytes and in 2-cell and 8-cell embryos. Notably, we show that ~22% of the oocyte genome is associated with broad H3K4me3 domains that are anti-correlated with DNA methylation. The H3K4me3 signal becomes confined to transcriptional-start-site regions in 2-cell embryos, concomitant with the onset of major zygotic genome activation. Active removal of broad H3K4me3 domains by the lysine demethylases KDM5A and KDM5B is required for normal zygotic genome activation and is essential for early embryo development. Our results provide insight into the onset of the developmental program in mouse embryos and demonstrate a role for broad H3K4me3 domains in MZT.

The importance of germline-inherited posttranslational histone modifications on priming early mammalian development is just emerging 1 – 4 . Histone H3 lysine 9 (H3K9) trimethylation is associated with heterochromatin and gene repression during cell-fate change 5 , while histone H3 lysine 4 (H3K4) trimethylation marks active gene promoters 6 . Mature oocytes are transcriptionally quiescent and possess remarkably broad domains of H3K4me3 (bdH3K4me3) 1 , 2 . It remains unknown as to which factors contribute to the maintenance of the bdH3K4me3 landscape. Lysine-specific demethylase 4A (KDM4A) demethylates H3K9me3 at promoters marked by H3K4me3 in actively transcribing somatic cells 7 . Here, we report that KDM4A-mediated H3K9me3 demethylation at bdH3K4me3 in oocytes is crucial for normal preimplantation development and zygotic genome activation (ZGA) after fertilization. Loss of KDM4A in oocytes causes aberrant H3K9me3 spreading over bdH3K4me3, resulting in insufficient transcriptional activation of genes, endogenous retroviral elements and long terminal repeat initiated chimeric transcripts during ZGA. The catalytic activity of KDM4A is essential for normal epigenetic reprogramming and preimplantation development. Hence, KDM4A plays a crucial role in preserving maternal epigenome integrity required for proper ZGA and transfer of developmental control to the embryo.

In most mammalian cells, DNA replication occurs once, and only once between cell divisions. Replication initiation is a highly regulated process with redundant mechanisms that prevent errant initiation events. In lower eukaryotes, replication is initiated from a defined consensus sequence, whereas a consensus sequence delineating mammalian origin of replication has not been identified. Here we show that 5-hydroxymethylcytosine (5hmC) is present at mammalian replication origins. Our data support the hypothesis that 5hmC has a role in cell cycle regulation. We show that 5hmC level is inversely proportional to proliferation; indeed, 5hmC negatively influences cell division by increasing the time a cell resides in G1. Our data suggest that 5hmC recruits replication-licensing factors, then is removed prior to or during origin firing. Later we propose that TET2, the enzyme catalyzing 5mC to 5hmC conversion, acts as barrier to rereplication. In a broader context, our results significantly advance the understating of 5hmC involvement in cell proliferation and disease states.

Neural stem/progenitor cells (NSPCs) persist in the mammalian brain throughout life and can be activated in response to the physiological and pathophysiological stimuli. Epigenetic reprogramming of NPSC represents a novel strategy for enhancing the intrinsic potential of the brain to regenerate after brain injury. Therefore, defining the epigenetic features of NSPCs is important for developing epigenetic therapies for targeted reprogramming of NSPCs to rescue neurologic function after injury. In this study, we aimed at defining different subtypes of NSPCs by individual histone methylations. We found the three histone marks, histone H3 lysine 4 trimethylation (H3K4me3), histone H3 lysine 27 trimethylation (H3K27me3), and histone H3 lysine 36 trimethylation (H3K36me3), to nicely and dynamically portray individual cell types during neurodevelopment. First, we found all three marks co-stained with NSPC marker SOX2 in mouse subventricular zone. Then, CD133, Id1, Mash1, and DCX immunostaining were used to define NSPC subtypes. Type E/B, B/C, and C/A cells showed high levels of H3K27me3, H3K36me3, and H3K4me3, respectively. Our results reveal defined histone methylations of NSPC subtypes supporting that epigenetic regulation is critical for neurogenesis and for maintaining NSPCs.

Early vertebrate embryogenesis is characterized by extensive post-transcriptional regulation during the maternal-to-zygotic transition. The N6-methyladenosine (m 6 A) modifications on mRNA have been shown to affect both translation and stability of transcripts. Here we investigate the m 6 A topology during early vertebrate embryogenesis and its association with polyadenylated mRNA levels. The majority (>70%) of maternal transcripts harbor m 6 A, and there is a substantial increase of m 6 A in the polyadenylated mRNA fraction between 0 and 2 hours post fertilization.Notably, we find strong associations between m 6 A, cytoplasmic polyadenylation and translational efficiency prior to zygotic genome activation (ZGA). Interestingly, the relationship between m 6 A and translation is strongest for peaks located in the 3'UTR, but not overlapping stop codons.Sequence analyses revealed enrichment of motifs for RNA binding proteins involved in translational regulation and RNA degradation. After ZGA, m 6 A seem to diminish the effect of miR-430 mediated degradation. The reported results improve our understanding of the combinatorial code behind post-transcriptional mRNA regulation during embryonic reprogramming and early differentiation. IntroductionPost-transcriptional chemical modifications of RNA alter their fate and are important for RNA function. The N6-methyladenosine (m 6 A) modification is the most abundant internal mRNA modification, and each mRNA contains on average three to five m 6 A modifications [1,2]. The fate and life-time of m 6 A containing mRNAs is partly regulated through specific interaction with the YTH-domain containing proteins. RNAs bound by YTHDF2 is transported to decay sites resulting in increased degradation rates [3], while mRNAs bound to YTHDF1 are more efficiently translated through interaction with the translation machinery [4]. In addition to these direct effects, m 6 A can change the secondary structure of transcripts and allow proteins to bind otherwise hidden sequence motifs [5,6].Zebrafish is a very attractive system to study embryogenesis and associated processes such as reprogramming and differentiation, including their associated transcriptomic and epigenetic blueprints. Although the zebrafish share embryonic features with other species, embryogenesis is remarkably fast compared with mammals ( Fig 1a). The egg is activated upon water contact, and after fertilization and the zygote period (1-cell; 0 to 0.75 hours post fertilization, hpf), the embryo start to divide rapidly during cleavage stages (0.75 to 2 hpf), reaching 64-cells before entering the blastula period (2.25 to 5.25 hpf). During this time span, zygotic genome activation (ZGA, ~3 hpf), occurs. At 4hpf ("sphere") there is robust expression of many genes important for further developmental progress [7,8]. The blastula period cells can form embryonic stem cells in culture, similar to the inner cell mass of mouse embryos [9, 10]. Thus, from fertilization until blastula, the germ cells have been reprogrammed to become pluripoten...