Spermatogonial stem cells (SSCs) are the adult stem cell population that upholds the sustainable male fertility. SSCs self-renew to maintain a pool of undifferentiated spermatogonia and differentiate to become sperm in response to developmental cues. The precise regulatory mechanisms governing the fate decision of SSCs between self-renewal and differentiation remain largely elusive. My first study in this dissertation investigated mitochondrial regulation of spermatogonial proliferation and differentiation. Mitochondria are dynamically changing organelles that alter their features through continuous cycles of fusion and fission processes that are collectively called mitochondrial dynamics. I found that during spermatogonial differentiation, pro-fusion factors (e.g., MFN1, OPA1) and pro-fission regulators (e.g., DRP1, FIS1) were both upregulated. In addition, I observed increased mitochondrial permeability transition pore (mPTP) opening and decreased mitochondrial membrane potential (MMP), accompanied with upregulated ROS level upon spermatogonial differentiation. Importantly, I demonstrated that enhanced mitochondrial dynamics promoted spermatogonial differentiation. My data also suggested that fusion regulator MFN1 increased spermatogonial differentiation through the regulation of metabolism, while fission factor DRP1 impacted spermatogonial differentiation via mPTP opening, thus providing novel information about functional mechanism that underlies mitochondrial regulation of spermatogonial development. My second study in this dissertation characterized the novel role of transcription factor FOXF1 in regulating spermatogonial proliferation and differentiation. I found that FOXF1 marked a subpopulation of undifferentiated spermatogonia in both neonatal and adult testes. In addition, I demonstrated that the deficiency of FOXF1 impaired SSCs activity. Using a spermatogonial differentiation platform, I revealed that spermatogonial differentiation were also affected by altered FOXF1 disturbance. Transcriptomic analysis further uncovered that FOXF1 might regulate the transition of SSC fate from quiescence to proliferation via cell cycle and metabolism. Taken together, my research findings will shed light on our understanding of critical regulation of mammalian spermatogonial proliferation and differentiation.
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