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Gonocyte-to-spermatogonia transition is a critical fate determination process to initiate sperm production throughout the lifecycle. However, the molecular dynamics of this process has not been fully elucidated mainly due to the asynchronized differentiation stages of neonatal germ cells. In this study, we employed single cell RNA sequencing analyses of P1.5–5.5 germ cells to clarify the temporal dynamics of gene expression during gonocyte-to-spermatogonia transition. The analyses identified transcriptional modules, one of which regulates spermatogonial gene network in neonatal germ cells. Among them, we identified  Dec2 , a bHLH-type transcription factor, as a transcriptional repressor for a spermatogonial differentiation factor  Sohlh1 . Deficiency of  Dec2  in mice induces significant reduction of undifferentiated spermatogonia, and transplantation assay using  Dec2 -depleted cells also demonstrated the impaired efficiency of engraftment, suggesting its role in maintaining spermatogonial stem cells (SSCs). Collectively, this study revealed the intrinsic role of a new SSC factor  Dec2 , which protects germ cells from inadequate differentiation during neonatal testis development.

Single cell RNA-sequencing identified Dec2 as a suppressive factor for spermatogonial differentiation by inhibiting Sohlh1 expression