No abstract

The arrest of meiotic prophase in mammalian oocytes within fully grown follicles is dependent on cyclic adenosine monophosphate (cAMP) regulation. A large part of cAMP is produced by the Gs-linked G-protein-coupled receptor (GPR) pathway. In the present study, we examined whether GPR3 is involved in the maintenance of meiotic arrest in porcine oocytes. Expression and distribution of GPR3 were examined by western blot and immunofluorescence microscopy, respectively. The results showed that GPR3 was expressed at various stages during porcine oocyte maturation. At the germinal vesicle (GV) stage, GPR3 displayed a maximal expression level, and its expression remained stable from pro-metaphase I (MI) to metaphase II (MII). Immunofluorescence staining showed that GPR3 was mainly distributed at the nuclear envelope during the GV stage and localized to the plasma membrane at pro-MI, MI and MII stages. RNA interference (RNAi) was used to knock down the GPR3 expression within oocytes. Injection of small interfering double-stranded RNA (siRNA) targeting GPR3 stimulated meiotic resumption of oocytes. On the other hand, overexpression of GPR3 inhibited meiotic maturation of porcine oocytes, which was caused by increase of cGMP and cAMP levels and inhibition of cyclin B accumulation. Furthermore, incubation of porcine oocytes with the GPR3 ligand sphingosylphosphorylcholine (SPC) inhibited oocyte maturation. We propose that GPR3 is required for maintenance of meiotic arrest in porcine oocytes through pathways involved in the regulation of cAMP and cGMP.

The Ska (spindle and kinetochore-associated) complex is composed of three proteins: Ska1, Ska2 and Ska3. It is required for stabilizing kinetochore-microtubule (KT-MT) interactions and silencing spindle checkpoint during mitosis. However, its roles in meiosis remain unclear. The present study was designed to investigate the localization and function of the Ska complex during mouse oocyte meiotic maturation. Our results showed that the localization and function of Ska complex in mouse oocyte meiosis differ in part from those in mitosis. Injection of low dose exogenous Myc-Ska mRNA showed that, instead of localizing to the kinetochores (KTs) and mediating KT-MT interactions from pro-metaphase to mid-anaphase stages as in mitosis, the members of the Ska complex were only localized on spindle microtubules from the Pro-MI to MII stages in mouse oocyte meiosis. Time-lapse live imaging analysis showed that knockdown of any member of the Ska complex by Morpholino injection into mouse oocytes resulted in spindle movement defects and enlarged polar bodies. Depletion of the whole Ska complex disrupted the stability of the anaphase spindle and influenced the extrusion of the first polar body. Taken together, these results show that the Ska complex plays an important role in meiotic spindle migration and anaphase spindle stability during mouse oocyte maturation.

No abstract

The mouse sperm genome is resistant to in vitro heat treatment, and embryos derived from heated sperm can support full-term embryonic development, but the blastocyst rate and implantation rate are lower compared to embryos derived from fresh sperm. In the present study, the patterns of DNA methylation, histone H4K12 (ACH4K12) acetylation, H3K9 trimethylation (H3K9-TriM), and H3K27 trimethylation (H3K27-TriM) in preimplantation embryos derived from 65 °C-heated sperm were investigated. Although no evident changes in global DNA methylation, histone H4K12 (ACH4K12) acetylation, and H3K9 trimethylation (H3K9-TriM) were found, significantly lower levels of H3K27-TriM, which was thought to be one of the reasons for low efficiency of mouse cloning, were found in the inner cell mass of heated-sperm derived blastocysts. Thus, defective modification of H3K27-TriM might contribute to compromised development of embryos derived from heated sperm.

UCHL5IP is one of the subunits of the haus complex, which is important for microtubule generation, spindle bipolarity and accurate chromosome segregation in Drosophila and human mitotic cells. In this study, the expression and localisation of UCHL5IP were explored, as well as its functions in mouse oocyte meiotic maturation. The results showed that the UCHL5IP protein level was consistent during oocyte maturation and it was localised to the meiotic spindle in MI and MII stages. Knockdown of UCHL5IP led to spindle defects, chromosome misalignment and disruption of γ-tubulin localisation in the spindle poles. These results suggest that UCHL5IP plays critical roles in spindle formation during mouse oocyte meiotic maturation.