The Transcriptional Cell Atlas of Testis Development in Sheep at Pre-Sexual Maturity.

Yi Wu,Shaohua Zhu,Jianye Li,Jianlin Han,Jianbin Liu,Tingting Guo,Guoyan Qiao,Hongchang Zhao,Weibo Sun,Chao Yuan,Zengkui Lu,Chune Niu,Yaojing Yue,Xue He,Mei Han,Bohui Yang

doi:10.3390/cimb44020033

Abstract

Sheep testes undergo a dramatic rate of development with structural changes during pre-sexual maturity, including the proliferation and maturation of somatic niche cells and the initiation of spermatogenesis. To explore this complex process, 12,843 testicular cells from three males at pre-sexual maturity (three-month-old) were sequenced using the 10× Genomics ChromiumTM single-cell RNA-seq (scRNA-seq) technology. Nine testicular somatic cell types (Sertoli cells, myoid cells, monocytes, macrophages, Leydig cells, dendritic cells, endothelial cells, smooth muscle cells, and leukocytes) and an unknown cell cluster were observed. In particular, five male germ cell types (including two types of undifferentiated spermatogonia (Apale and Adark), primary spermatocytes, secondary spermatocytes, and sperm cells) were identified. Interestingly, Apale and Adark were found to be two distinct states of undifferentiated spermatogonia. Further analysis identified specific marker genes, including UCHL1, DDX4, SOHLH1, KITLG, and PCNA, in the germ cells at different states of differentiation. The study revealed significant changes in germline stem cells at pre-sexual maturation, paving the way to explore the candidate factors and pathways for the regulation of germ and somatic cells, and to provide us with opportunities for the establishment of livestock stem cell breeding programs.

Highlights

Sexual maturity refers to the period after puberty when the male’s body and reproductive organs are further developed, reproductive function is improved, and normal fertility is achieved [1]
The expressions of known marker genes in the testicular tissue and the tissue derived RNA were evaluated after PCR amplification, as displayed in Supplementary Figure S2, including the reference genes of GAPDH, VWF, NOTCH3, IGF1, ACTA2, SOX9, CD83, PENK, S100B, BEX4, PEGFRB, NR6A1, ID4, CDH11, DAZL, BCAT2, PCNA, KITLG, DDX4, TOP2A, UCHL1, and SOHLH1
The results indicated that CDH11, UCHL1, and DDX4 were highly expressed in undifferentiated spermatogonia, while there was no expression in spermatocyte or spermatids

Summary

Introduction

Sexual maturity refers to the period after puberty when the male’s body and reproductive organs are further developed, reproductive function is improved, and normal fertility is achieved [1]. During this period, the testes are involved in major physiological changes, including a large increase in their volume and the initiation of somatic cell proliferation/maturation and spermatogenesis [2]. Single-cell RNA-seq (scRNA-seq) technology has a high cell capture rate, superior sensitivity, and data accuracy, which better reflect cellular heterogeneity [14]

Methods

Results

Conclusion