Gene Regulatory Networks Orchestrating Oocyte Fate Bifurcation in Primordial Follicles Revealed by Single-Cell Transcriptomics

The first wave of primordial follicle activation occurs around postnatal day 2.5 (P2.5), with the remaining follicles entering a dormant state to ensure a continuous supply of fertilizable oocytes. However, the molecular mechanisms underlying this critical fate bifurcation remain poorly understood. Here, we employ SMART-seq2 to profile transcriptional dynamics in individual perinatal female mouse germ cells across three developmental stages: cysts at embryonic day 17.5 (E17.5), primordial follicles at P2.5, and primary follicles at P6.5. Unsupervised clustering and trajectory inference reveal two distinct transcriptional states in P2.5 oocytes, corresponding to dormant and activated subpopulations. Activated oocytes exhibit upregulation of genes linked to PI3K-Akt/mTORC1 signaling, extracellular matrix (ECM) disassembly, and oocyte maturation, whereas marker genes of dormant oocytes are enriched in oxidative stress response and DNA repair pathways. Transcriptional regulatory network reconstruction based on SCENIC inference identifies key transcription factors (e.g., Bhlhe41, Tcf3/12) orchestrating the fate bifurcation. Notably, dormant oocytes form a stable transcriptional state distinct from the continuous activation and development trajectory. Our findings provide a comprehensive landscape of gene expression and regulatory networks correlated to fate determination of P2.5 primordial follicles, paving the way for advanced therapeutic strategies for related diseases and improved techniques for in vitro follicle activation.