Oocyte morphology and transcriptional silencing in relation to chromatin remodeling during the final phases of bovine oocyte growth

Abstract

The complexity of the events which orchestrate mammalian oocyte growth and ultimate acquisition of developmental competence is still unclear and under continuous investigation. Starting from the observation that germinal vesicle (GV) bovine oocytes exhibit different patterns of chromatin configuration (from GV0 to GV3), the present study aimed at analyzing the morphology of the nuclear and the cytoplasmic compartments and to determine the total transcriptional activity of immature oocytes sorted on the basis of their chromatin configuration. The oocyte morphology was analyzed by light microscopy (LM) on semi-thin sections and transmission electron microscopy, and the global transcriptional activity was analyzed by H(3)-Uridine incorporation and subsequent autoradiography on semi-thin sections. LM confirmed the increase in chromatin condensation from GV0 to GV3. Ultrastructurally, the nucleolus was fibrillo granular at GV0 while the other stages displayed an electron-dense fibrillar sphere with the remnant of a fibrillar center on the surface. Organelles were dispersed in the cytoplasm at GV0 while at GV1 and GV2 most of them were homogenously distributed in the oocyte cortex. At GV3 most organelles were found in clusters in the oocyte cortex. Typical features of completion of the oocyte growth phase, like undulation of the nuclear envelope and reduction of the size of Golgi complex were found at GV2 and GV3. Moreover, GV3 oocytes presented cortical granules that displayed varying degrees of degeneration. Autoradiographic labeling showed that oocytes with GV0 configuration exhibited high level of RNA synthesis, GV1 and GV2 stages showed a remarkable decrease of transcription, and the acquisition of GV3 configuration was associated with global repression of transcriptional activity. Our findings suggest a temporal relationship between the chromatin remodeling process and the main morpho-functional events that characterize the final growth phase in bovine oocyte.

Role of gap junction-mediated communications in regulating large-scale chromatin configuration remodeling and embryonic developmental competence acquisition in fully grown bovine oocyte

“…This remodeling process has been related to the progressive increase of embryonic developmental competence [3,8]. However, the mechanisms and factors controlling large-scale chromatin configuration changes have remained elusive.…”

RNA Processing During Early Embryogenesis: Managing Storage, Utilisation and Destruction

“…In bovine oocyte maturation, the terms GV0-GV3 are predominantly used (Lodde et al 2008) though SN and Non SN configurations are also referred to in literature (Tan et al 2009). In GV0 oocytes, there is much transcription in the nucleolus and a number of morphological differences exist that distinguish the bovine oocyte from later GV stages that include erected microvilli, small clusters of immature mitochondria, few ooplasmic vesicles, the presence of a golgi apparatus, and scattered cortical granules (Lodde et al 2008).…”

Localization of DNA methyltransferase-1 during oocyte differentiation, in vitro maturation and early embryonic development in cow

“…Oocytes collected from early and middle antral follicles were freed of cumulus cells and divided according to their chromatin configuration as previously described (Lodde et al, 2007;Lodde et al, 2008). Briefly, denuded oocytes (DOs) were obtained by mechanically removing of cumulus cells by the use of vortex (2 min, 35Htz) in M199 supplemented with HEPES 20 mM and with 5% of calf serum (HM199).…”

“…Our data indicate that DNA methylation is already established in GV0 stage oocytes and slightly increases thereafter with chromatin condensation in GV1 oocytes, which in turn seems to mark the achievement of a stable level of global DNA methylation as no further differences where observed between GV1, GV2 and GV3 stages oocytes. The increased DNA methylation from GV0 to GV1 stage oocytes correlates with the decrease of transcriptional activity that was observed during this transition (Lodde et al, 2008). In fact, a growing body of evidence indicates that the chromatin structure together with DNA methylation mutually reinforce the transcriptional repressive state of DNA (Bird, 2002;Fuks et al, 2003) their limited competences as it is becoming evident that genome wide epigenetic modifications must be set in place during gametogenesis in order to allow a correct developmental program (Albertini et al, 2003;Bao et al, 2000;De La Fuente, 2006;Li, 2002).…”

“…Our previous studies indicated that the structural chromatin transitions in the oocyte nucleus are temporally related to the main morphological and functional events that characterize the final growth phase in bovine oocyte and thus representing a marker of oocyte differentiation. In particular, the transition from the GV0 to the GV1 stage has been related to the global transcription silencing and to the acquisition of meiotic competence, while the transition from the GV1 stage to higher degrees of condensation in GV2 and GV3 stage oocytes marks the acquisition of a full embryonic developmental capability (Lodde et al, 2007;Lodde et al, 2008). Although DNA methylation patterns in bovine oocytes and early embryos have been already documented in previous reports (Bourc 'his et al, 2001;Dean et al, 2001;Kang et al, 2001;Maalouf et al, 2008), the present study analyzes for the first time this epigenetic mark according to the chromatin configuration changes.…”