Genome organization and epigenetic marks in mouse germinal vesicle oocytes

Abstract

During the final step of oogenesis, the oocyte nucleus is subject to large-scale modifications that correlate with transcriptional silencing. While oocytes with dense chromatin around the nucleolus are silent (SN, surrounded nucleolus), oocytes with uncondensed chromatin (NSN, non-surrounded nucleolus) are transcriptionally active. It is believed that epigenetic mechanisms that participate in gene expression regulation could play a role in this event. In this context, we examined the behaviour of heterochromatin and related histone modifications during the NSN to SN transition by immunostaining. Using fluorescent in situ hybridization on three dimensionalpreserved nuclei (3D-FISH), we also studied the distribution of centromeric, pericentromeric and ribosomal (rDNA) sequences in relation to the nucleolus (also called the nucleolus-like body, NLB). We observed that in NSN-type oocytes, pericentromeric heterochromatin is aggregated within chromocenters. In SN-type oocytes, pericentromeric heterochromatin and centromeres form a discontinuous ring around the NLB. rDNA sequences, which initially present a pearl necklace structure, gather together in seven highly condensed foci at the NLB periphery. H3K9me3 and H4K20me3 heterochromatin marks clearly label chromocenters, whereas H3K4me3 and H4K5ac are totally excluded from heterochromatin regions, even in the very compact SN-nuclei. Remarkably, H3K27me3 displays an intermediate behavior. It appears that GV oocyte nuclei exhibit a specific epigenetic landscape. Histone modifications, related to both active and repressive chromatin structures, seem to follow the large-scale chromatin movements that occur during the NSN to SN transition. We also demonstrate that, while heterochromatin regions re-localize around the NLB, rDNA sequences adopt a highly compact structure in SN-type oocytes.

Localisation of RNAs and proteins in nucleolar precursor bodies of early mouse embryos

“…NPBs, like NLBs, are comprised of a tightly packed fibrous material, lack the typical nucleolar tripartite organisation and are transcriptionally silent despite their association with ribosomal genes (rDNA; Fakan and Odartchenko 1980;Tesarík et al 1987;Fléchon and Kopecný 1998;Laurincik et al 2000;Romanova et al 2006b;Bonnet-Garnier et al 2012). The morphology of NPBs remains unchanged until the main resumption of rDNA transcription commences in late two-cell stage in the mouse (Engel et al 1977;Zatsepina et al 2003;Bogolyubova 2011) and in more advanced developmental stages in embryos of other mammalian species (Braude et al 1988;Hyttel et al 2000;Bjerregaard et al 2004;Graf et al 2014).…”

Restraint Stress on Female Mice Diminishes the Developmental Potential of Oocytes: Roles of Chromatin Configuration and Histone Modification in Germinal Vesicle Stage Oocytes1

“…Although the nonsurrounded nucleolus (NSN) oocytes have less condensed chromatin, which does not form a heterochromatin rim around the nucleolus, the SN (surrounded nucleolus) oocytes show condensed chromatin that is particularly confined around the nucleolus [1][2][3][4]. Generally, oocyte competence appears to be influenced primarily by epigenetic factors that control overall gene expression while significantly modifying GV chromatin configuration [5][6][7][8]. For example, fully grown GV oocytes must end an NSN configuration before gaining full meiotic competence, and they must take on an SN configuration and stop gene transcription before having the capacity to sustain blastocyst formation [6].…”

Visualization of the spatial arrangement of nuclear organization using three-dimensional fluorescence <i>in situ</i> hybridization in early mouse embryos: A new “EASI-FISH chamber glass” for mammalian embryos

“…To date, 3D-FISH analysis in oocytes and embryos has been confined to a few reports [3, 4, 10,11,12, 24]. The reasons for the limitations of the studies were the unique morphological and cytological properties of the embryos.…”

“…However, 3D-FISH in oocytes and early embryos in mammals has been confined to reports concerning the repeated sequences in centromeres and telomeres, as well as rDNA [3, 4, 11, 12], because of the unique properties attributable to a large spherical cell with a diameter of 100 µm, which interferes with the permeation/elution of probes because of the large amount of ooplasm/cytoplasm and requires fine manipulation skill using micro-capillaries under microscopes throughout the process of FISH. In this study, we devised a new “EASI-FISH chamber glass” for 3D-FISH in early embryos, which enabled convenient and reproducible anchoring of the embryo onto the coverslip, while retaining its steric structure throughout the FISH process.…”