Establishing the Oocyte Proteome at the Primordial Follicle Stage and Cisplatin-induced Shifts through in situ APEX2-driven Biotinylation

The number and quality of primordial follicles play a decisive role in female fertility. The high sensitivity of oocytes in primordial follicles to chemotherapeutics makes antitumor therapy extremely prone to premature ovarian insufficiency and infertility; however, the specific mechanism remains unclear. A systematic proteomic profile of oocytes in primordial follicles is urgently needed to support basic and clinical research, which has been hindered by the rarity of oocyte samples and the technical challenges associated with isolating oocytes from primordial follicles. In this study, utilizing in vivo protein labeling by engineered ascorbate peroxidase (APEX) fluorescent mice, we identified 641 proteins and 2020 gene transcripts in primordial follicular oocytes, from which the abundance of several proteins involved in processes associated with DNA damage repair and histone modification changed after ovary exposure to the chemotherapeutic cisplatin. Notably, a histone modifier, EZH2, was induced by cisplatin in oocytes, and the simultaneous application of its inhibitor, GSK126, partially relieved cisplatin-induced oocyte developmental defects. Our study provides the first description of the primordial follicular oocyte proteome and further illustrates the dynamic shifts in protein abundance associated with chemotherapeutic agents, laying the foundation for further development of effective and targeted drugs to protect and prolong female fertility. Overall design: The biotinylated RNAs from APEXflox/+;Gdf9-Cre mice were subjected to RNA-seq using the Smart-seq2 method with minor modifications.