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

Yoshinori Makino,Haruhiko Akiyama,Niels H Jensen,Yuki Okada,Katsuhiko Shirahige,Naoko Yokota,Moritz J Rossner

doi:10.1038/s41598-019-42578-z

Abstract

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.

Highlights

Spermatogenesis is a process to produce spermatozoa almost throughout the lifecycle, and sustained by spermatogonial stem cells (SSCs) that can both self-renew and differentiate[1,2,3]
To more precisely understand gonocyte-to-spermatogonia transition from the perspective of cellular heterogeneity and temporally regulated gene expression patterns, male germ cells were collected from P1.5, P3.5, and P5.5 testes and subjected to scRNA-seq
The molecular basis of gonocyte-to-spermatogonia transition in neonatal testis has been an important and abstruse question due to the heterogeneous cell population occurring during this period

Summary

Introduction

Spermatogenesis is a process to produce spermatozoa almost throughout the lifecycle, and sustained by spermatogonial stem cells (SSCs) that can both self-renew and differentiate[1,2,3]. SSC/undifferentiated to differentiating state of spermatogonia are characterized by the transition from higher expressions of Id4, Gfra[1], Etv[5], and Lhx[1] to Sohlh1/2, Kit, and Star[8] as numbers of former studies have been demonstrated by immunostaining, RT-PCR, etc.[16,17,18,19,20,21,22,23]. This transition is physiologically induced by retinoic acid (RA) stimulation, the underlying molecular pathway has not been fully elucidated. We experimentally demonstrated that Dec[2] plays an inhibitory role in spermatogonial differentiation in neonatal germ cells by suppressing Sohlh[1] expression

Methods

Results

Conclusion