DNA repair protein RAD51 homolog 1 (RAD51) plays a central role in homologous recombination (HR) repair of DNA breaks. HR depends on the formation of a RAD51 recombinase filament that facilitates strand invasion. However, the role of RAD51 during porcine oocyte maturation is unknown. The objective of this study was to investigate the expression and function of RAD51 during porcine oocyte maturation in vitro. RAD51 was mainly localized to the nucleus at the germinal vesicle (GV) stage, and was widely distributed in the cytoplasm between the GV breakdown (GVBD) and metaphase II stage. DNA damage induced by etoposide was accompanied by the formation of RAD51 foci that were colocalized with γH2AX. Inhibition of RAD51 increased DNA damage and induced metaphase I arrest along with spindle defects, chromosomal misalignment, and abnormal spindle assembly checkpoint (SAC) activity. Inhibition of RAD51 also increased ROS levels and led to an abnormal mitochondrial distribution. Our results indicate that RAD51 plays a critical role in maintaining chromosome integrity and mitochondrial activity during porcine oocyte maturation.

Imperatorin (IMP) exhibits a variety of pharmacological properties, including antioxidant, anti-inflammatory, antibacterial, anti-cancer, and anti-hypertension activities. However, its effects on animal reproduction systems, especially oocyte development, maturation, and aging are not yet clear. In this study, the effects of IMP on oocyte development and aging as well as the underlying molecular mechanisms were explored. Oocytes were cultured for an additional 24 h for aging. Results revealed that the blastocyst formation and hatching rates of embryos, which were parthenogenetically activated aged oocytes, were significantly increased with IMP treatment (40 μM). Simultaneously, well-distributed cortical granules but no significant difference in zona pellucida hardness were observed after IMP treatment. During this stage, intracellular reactive oxygen species, apoptosis, and autophagy levels were decreased, while mitochondrial membrane potential, glutathione level, and activity of superoxide dismutase and catalase were increased. IMP-treated aged oocytes also showed significantly higher expression of MOS, CCNB1, BMP15, and GDF9 than non-IMP-treated aged oocytes although their levels were still lower than those in the fresh oocytes. These results suggest that IMP can effectively ameliorate the quality of aged porcine oocytes by reducing oxidative stress and protecting mitochondrial function.

Homologous recombination (HR) plays a critical role in facilitating replication fork progression when the polymerase complex encounters a blocking DNA lesion, and it also serves as the primary mechanism for error-free DNA repair of double-stranded breaks. DNA repair protein RAD51 homolog 1 (RAD51) plays a central role in HR. However, the role of RAD51 during porcine early embryo development is unknown. In the present study, we examined whether RAD51 is involved in the regulation of early embryonic development of porcine parthenotes. We found that inhibition of RAD51 delayed cleavage and ceased development before the blastocyst stage. Disrupting RAD51 activity with RNAi or an inhibitor induces sustained DNA damage, as demonstrated by the formation of distinct γH2AX foci in nuclei of four-cell embryos. Inhibiting RAD51 triggers a DNA damage checkpoint by activating the ataxia telangiectasia mutated (ATM)–p53–p21 pathway. Furthermore, RAD51 inhibition caused apoptosis, reactive oxygen species accumulation, abnormal mitochondrial distribution and decreased pluripotent gene expression in blastocysts. Thus, our results indicate that RAD51 is required for proper porcine parthenogenetic activation (PA) embryo development.

Actin-interacting protein 1 (AIP1), also known as WD repeat-containing protein 1 (WDR1), is ubiquitous in eukaryotic organisms, and it plays critical roles in the dynamic reorganization of the actin cytoskeleton. However, the biological function and mechanism of AIP1 in mammalian oocyte maturation is still largely unclear. In this study, we demonstrated that AIP1 boosts ADF/Cofilin activity in mouse oocytes. AIP1 is primarily distributed around the spindle region during oocyte maturation, and its depletion impairs meiotic spindle migration and asymmetric division. The knockdown of AIP1 resulted in the gathering of a large number of actin-positive patches around the spindle region. This effect was reduced by human AIP1 (hAIP1) or Cofilin (S3A) expression. AIP1 knockdown also reduced the phosphorylation of Cofilin near the spindle, indicating that AIP1 interacts with ADF/Cofilin-decorated actin filaments and enhances filament disassembly. Moreover, the deletion of AIP1 disrupts Cofilin localization in metaphase I (MI) and induces cytokinesis defects in metaphase II (MII). Taken together, our results provide evidence that AIP1 promotes actin dynamics and cytokinesis via Cofilin in the gametes of female mice.