Spatio-temporal landscape of mouse epididymal cells and specific mitochondria-rich segments defined by large-scale single-cell RNA-seq

Abstract

Spermatozoa acquire their fertilizing ability and forward motility during epididymal transit, suggesting the importance of the epididymis. Although the cell atlas of the epididymis was reported recently, the heterogeneity of the cells and the gene expression profile in the epididymal tube are still largely unknown. Considering single-cell RNA sequencing results, we thoroughly studied the cell composition, spatio-temporal differences in differentially expressed genes (DEGs) in epididymal segments and mitochondria throughout the epididymis with sufficient cell numbers. In total, 40,623 cells were detected and further clustered into 8 identified cell populations. Focused analyses revealed the subpopulations of principal cells, basal cells, clear/narrow cells, and halo/T cells. Notably, two subtypes of principal cells, the Prc7 and Prc8 subpopulations were enriched as stereocilia-like cells according to GO analysis. Further analysis demonstrated the spatially specific pattern of the DEGs in each cell cluster. Unexpectedly, the abundance of mitochondria and mitochondrial transcription (MT) was found to be higher in the corpus and cauda epididymis than in the caput epididymis by scRNA-seq, immunostaining, and qPCR validation. In addition, the spatio-temporal profile of the DEGs from the P42 and P56 epididymis, including transiting spermatozoa, was depicted. Overall, our study presented the single-cell transcriptome atlas of the mouse epididymis and revealed the novel distribution pattern of mitochondria and key genes that may be linked to sperm functionalities in the first wave and subsequent wave of sperm, providing a roadmap to be emulated in efforts to achieve sperm maturation regulation in the epididymis.

Anatomical Transcriptome Atlas of the Male Mouse Reproductive System During Aging

“…We identified a series of region-specific genes of male mouse reproductive tract ( Figures 2B,C ), which were in broad agreement with those of previous RNA-seq and microarray analyses of murine transcriptome profiles, such as ADAM metallopeptidase domain 28 ( Adam28 ) in initial segment, Transmembrane Epididymal Protein 1 ( Teddm1b ) in caput, and serine peptidase inhibitor, Kunitz type 3 ( Spint3 ) in corpus epididymis ( Johnston et al, 2005 ; Shi et al, 2021 ) (reviewed in Supplementary Table S4 ). GO analysis revealed the functional characteristics of the region-specific genes ( Figure 2D ; Supplementary Table S5 ).…”

Single Cell Transcriptome Sequencing of Zebrafish Testis Revealed Novel Spermatogenesis Marker Genes and Stronger Leydig-Germ Cell Paracrine Interactions

“…In order to study the cell-cell communication among cell populations, the Ligand-Receptor (LR) pairs were analyzed, which followed the procedure with minor modifications described in one recently published paper ( Shi et al, 2021 ) about LR studies in mouse. In detail, CellTalkDB ( Shao et al, 2020 ) was selected as the database for LR analyses, which contained 3,398 manually curated human LR pairs.…”

Calcium Homeostasis in the Epididymal Microenvironment: Is Extracellular Calcium a Cofactor for Matrix Gla Protein-Dependent Scavenging Regulated by Vitamins

“…Epididymal function involves a network of complex regulatory mechanisms that are not yet fully understood. Using state-of-the-art tools, such as single-cell omics and systems biology analyses, recent studies have revealed the complexity of cell biology in epididymal function ( de Lima et al, 2021 ; Rinaldi et al, 2020 ; Shi et al, 2021 ). A diagram of a basic model for the epithelial function, regulation of luminal microenvironment, and cell-cell interactions in the epididymis is presented in Figure 2 .…”