Abstract
High-quality oocyte production strategies play an important role in animal-assisted reproductive biotechnologies, and rapamycin (Rap) has been commonly used to increase the development potential of mammalian oocytes. The purpose of this study is to evaluate the effects and possible molecular mechanisms of rap on the maturation of yak oocytes. Different concentrations of Rap were supplemented during in vitro maturation (IVM) of yak oocytes. The maturation rates of oocytes and development rates of parthenogenetically activated embryos were assessed. The levels of 17β-estradiol (E2) were detected via ELISA, and the expression of autophagy-related factors, steroidogenic enzymes, and HIF-1α was detected via qRT-PCR, western blotting, and fluorescence microscopy, respectively. In addition, the impacts of E2 and HIF-1α on Rap-mediated oocyte autophagy were investigated by investigating the activities of E2 and HIF-1α. Our results showed that 0.1 nM Rap substantially enhanced the developmental ability of yak oocytes. In this group, the levels of E2, CYP19A1, CYP17A1, and autophagy-related factors were also significantly increased, and the expression of ATG5 and BECN1 in subsequent embryos was also increased. Further analysis revealed that Rap tends to enhance the development competence of yak oocytes and that the levels of autophagy-related factors are reduced when the activity of E2 or HIF-1α is inhibited. Furthermore, the levels of E2, CYP19A1, and CYP17A1 were downregulated when the activity of HIF-1α was inhibited, and the levels of HIF-1α were also significantly reduced by the estrogen receptor antagonist G15. Nevertheless, the levels of CYP11A1 mRNA in mature yak COCs were not significantly different among these groups, a phenomenon which implies that the levels of E2 were not correlated with the CYP11A1 content in yak COCs. There was an increasing tendency for the development competence of yak oocytes at the optimum concentration of Rap during IVM. The potential underlying mechanism is that Rap can activate autophagy and upregulate the levels of E2 and HIF-1α in mature oocytes. Additionally, the levels of both E2 and HIF-1α are regulated by each other and involve Rap-regulated autophagy in oocytes.