Single-cell RNA-sequencing reveals the dynamic process and novel markers in porcine spermatogenesis

Abstract

Background Spermatogenesis is the process by which male gametes are formed from spermatogonial stem cells and it is essential for the reliable transmission of genetic information between generations. To date, the dynamic transcriptional changes of defined populations of male germ cells in pigs have not been reported. Results To characterize the atlas of porcine spermatogenesis, we profiled the transcriptomes of ~ 16,966 testicular cells from a 150-day-old pig testis through single-cell RNA-sequencing (scRNA-seq). The scRNA-seq analysis identified spermatogonia, spermatocytes, spermatids and three somatic cell types in porcine testes. The functional enrichment analysis demonstrated that these cell types played diverse roles in porcine spermatogenesis. The accuracy of the defined porcine germ cell types was further validated by comparing the data from scRNA-seq with those from bulk RNA-seq. Since we delineated four distinct spermatogonial subsets, we further identified CD99 and PODXL2 as novel cell surface markers for undifferentiated and differentiating spermatogonia, respectively. Conclusions The present study has for the first time analyzed the transcriptome of male germ cells and somatic cells in porcine testes through scRNA-seq. Four subsets of spermatogonia were identified and two novel cell surface markers were discovered, which would be helpful for studies on spermatogonial differentiation in pigs. The datasets offer valuable information on porcine spermatogenesis, and pave the way for identification of key molecular markers involved in development of male germ cells.

Single-cell RNA sequencing uncovers dynamic roadmap and cell-cell communication during buffalo spermatogenesis

“…In mP spermatocytes, the central element deposition-related marker HORMAD2 and the pseudoautosomal region-related marker REC114 66 , 67 , 68 were substantially expressed. In lP spermatocytes, the tubulin-binding and microtubule motor activity-related genes NME5 and NME8 26 , 32 , 65 were expressed. The D spermatocytes express aurora kinase A/B ( AURKA and AURKB ).…”

“… 30 Furthermore, single-cell transcript levels of various cell types during spermatogenesis in sheep and pigs have been demonstrated. 31 , 32 Hence, the development patterns and gene expression of spermatogenic cells during spermatogenesis have both similarities and differences among species. 33 …”

Sertoli cell survival and barrier function are regulated by miR-181c/d-Pafah1b1 axis during mammalian spermatogenesis

“…Platelet-activating factor acetylhydrolase 1B subunit 1 (PAFAH1B1) (also known as Lissencephaly-1 (LIS1)) contains an N-terminal Lish domain and seven WD40 repeats at the C-terminal [20,21]. Immunohistochemical staining of mouse testicular tissues showed PAFAH1B1 is localized in spermatogenic cells and Sertoli cells [22], and single-cell RNA-sequencing data effectively validates that PAFAH1B1 is expressed in germ cells, Sertoli cells, Leydig cells, and other cells of pig [23] and mouse [24] testis. Deletion of Pafah1b1 in mice results in the failure of spermatids to form acrosomes and germ cell apoptosis [25,26].…”

Sertoli cell survival and barrier function are regulated by miR-181c/d-Pafah1b1 axis during mammalian spermatogenesis

“…Platelet-activating factor acetylhydrolase 1B subunit 1 (PAFAH1B1) (also known as Lissencephaly-1 (LIS1)) contains an N-terminal Lish domain and seven WD40 repeats at the C-terminal [20,21]. Immunohistochemical staining of mouse testicular tissues showed PAFAH1B1 is localized in spermatogenic cells and Sertoli cells [22], and single-cell RNA-sequencing data effectively validates that PAFAH1B1 is expressed in germ cells, Sertoli cells, Leydig cells and other cells of pig [23] and mouse [24] testis. Deletion of Pafah1b1 in mice results in the failure of spermatids to form acrosomes and germ cell apoptosis [25,26].…”