Single-cell RNA-Seq Resolves Cellular Heterogeneity and Transcriptional Dynamics during Spermatogonia Stem Cells Establishment and Differentiation

Abstract

The transition of gonocytes to spermatogonia and subsequent differentiation provide the foundation of spermatogenesis. However, systematic understanding on the cellular and molecular basis of this process is still limited, mainly impeded by the asynchrony in development and the lack of stage-specific markers. Using single-cell RNA sequencing on Oct4-GFP+/KIT- cells isolated from PND5.5 mice, we dissected the cellular heterogeneity and established molecular regulations. We demonstrated that gonocyte-spermatogonial transition was characterized by gene expression change related to apoptosis, cell cycle progression, and regulation of migration processes. Pseudotime analysis reconstructed developmental dynamics of the spermatogonial populations and unraveled sequential cellular and molecular transitions. We also identified CD87 as a neonatal stem cell marker which are potentially involved in the intial establishment of SSC pool. Lastly, we uncovered an unexpected subpopulation of spermatogonia primed to differentiation within the undifferentiated compartment, which is characterized by the lack of self-renewal genes and enhanced Oct4 expression and retinoic acid signaling response. Our study thus provides a novel understanding of cellular and molecular changes during spermatogonial establishment.