Title : Dissecting Cell Lineage Specification and Sex Fate Determination in Gonadal Somatic Cells Using Single-Cell Transcriptomics.

Publication date : 03. 2019

Authors : I Stévant, F Kühne, A Greenfield, MC Chaboissier, ET Dermitzakis, S Nef

Sex determination is a unique process that allows the study of multipotent progenitors and their acquisition of sex-specific fates during differentiation of the gonad into a testis or an ovary. Using time series single-cell RNA sequencing (scRNA-seq) on ovarian Nr5a1-GFP

Title : The Mammalian Spermatogenesis Single-Cell Transcriptome, from Spermatogonial Stem Cells to Spermatids.

Publication date : 11. 2018

Authors : BP Hermann, K Cheng, A Singh, L Roa-De La Cruz, KN Mutoji, IC Chen, H Gildersleeve, JD Lehle, M Mayo, B Westernströer, NC Law, MJ Oatley, EK Velte, BA Niedenberger, D Fritze, S Silber, CB Geyer, JM Oatley, JR McCarrey

Spermatogenesis is a complex and dynamic cellular differentiation process critical to male reproduction and sustained by spermatogonial stem cells (SSCs). Although patterns of gene expression have been described for aggregates of certain spermatogenic cell types, the full continuum of gene expression patterns underlying ongoing spermatogenesis in steady state was previously unclear. Here, we catalog single-cell transcriptomes for >62,000 individual spermatogenic cells from immature (postnatal day 6) and adult male mice and adult men. This allowed us to resolve SSC and progenitor spermatogonia, elucidate the full range of gene expression changes during male meiosis and spermiogenesis, and derive unique gene expression signatures for multiple mouse and human spermatogenic cell types and/or subtypes. These transcriptome datasets provide an information-rich resource for studies of SSCs, male meiosis, testicular cancer, male infertility, or contraceptive development, as well as a gene expression roadmap to be emulated in efforts to achieve spermatogenesis in vitro.

Title : The Mammalian Spermatogenesis Single-Cell Transcriptome, from Spermatogonial Stem Cells to Spermatids.

Publication date : 11. 2018

Authors : BP Hermann, K Cheng, A Singh, L Roa-De La Cruz, KN Mutoji, IC Chen, H Gildersleeve, JD Lehle, M Mayo, B Westernströer, NC Law, MJ Oatley, EK Velte, BA Niedenberger, D Fritze, S Silber, CB Geyer, JM Oatley, JR McCarrey

Spermatogenesis is a complex and dynamic cellular differentiation process critical to male reproduction and sustained by spermatogonial stem cells (SSCs). Although patterns of gene expression have been described for aggregates of certain spermatogenic cell types, the full continuum of gene expression patterns underlying ongoing spermatogenesis in steady state was previously unclear. Here, we catalog single-cell transcriptomes for >62,000 individual spermatogenic cells from immature (postnatal day 6) and adult male mice and adult men. This allowed us to resolve SSC and progenitor spermatogonia, elucidate the full range of gene expression changes during male meiosis and spermiogenesis, and derive unique gene expression signatures for multiple mouse and human spermatogenic cell types and/or subtypes. These transcriptome datasets provide an information-rich resource for studies of SSCs, male meiosis, testicular cancer, male infertility, or contraceptive development, as well as a gene expression roadmap to be emulated in efforts to achieve spermatogenesis in vitro.

Title : Single-Cell RNA Sequencing Analysis Reveals Sequential Cell Fate Transition during Human Spermatogenesis.

Publication date : 10. 2018

Authors : M Wang, X Liu, G Chang, Y Chen, G An, L Yan, S Gao, Y Xu, Y Cui, J Dong, Y Chen, X Fan, Y Hu, K Song, X Zhu, Y Gao, Z Yao, S Bian, Y Hou, J Lu, R Wang, Y Fan, Y Lian, W Tang, Y Wang, J Liu, L Zhao, L Wang, Z Liu, R Yuan, Y Shi, B Hu, X Ren, F Tang, XY Zhao, J Qiao

Spermatogenesis generates mature male gametes and is critical for the proper transmission of genetic information between generations. However, the developmental landscapes of human spermatogenesis remain unknown. Here, we performed single-cell RNA sequencing (scRNA-seq) analysis for 2,854 testicular cells from donors with normal spermatogenesis and 174 testicular cells from one nonobstructive azoospermia (NOA) donor. A hierarchical model was established, which was characterized by the sequential and stepwise development of three spermatogonia subtypes, seven spermatocyte subtypes, and four spermatid subtypes. Further analysis identified several stage-specific marker genes of human germ cells, such as HMGA1, PIWIL4, TEX29, SCML1, and CCDC112. Moreover, we identified altered gene expression patterns in the testicular somatic cells of one NOA patient via scRNA-seq analysis, paving the way for further diagnosis of male infertility. Our work allows for the reconstruction of transcriptional programs inherent to sequential cell fate transition during human spermatogenesis and has implications for deciphering male-related reproductive disorders.

Title : A Comprehensive Roadmap of Murine Spermatogenesis Defined by Single-Cell RNA-Seq.

Publication date : 09. 2018

Authors : CD Green, Q Ma, GL Manske, AN Shami, X Zheng, S Marini, L Moritz, C Sultan, SJ Gurczynski, BB Moore, MD Tallquist, JZ Li, SS Hammoud

Spermatogenesis requires intricate interactions between the germline and somatic cells. Within a given cross section of a seminiferous tubule, multiple germ and somatic cell types co-occur. This cellular heterogeneity has made it difficult to profile distinct cell types at different stages of development. To address this challenge, we collected single-cell RNA sequencing data from ∼35,000 cells from the adult mouse testis and identified all known germ and somatic cells, as well as two unexpected somatic cell types. Our analysis revealed a continuous developmental trajectory of germ cells from spermatogonia to spermatids and identified candidate transcriptional regulators at several transition points during differentiation. Focused analyses delineated four subtypes of spermatogonia and nine subtypes of Sertoli cells; the latter linked to histologically defined developmental stages over the seminiferous epithelial cycle. Overall, this high-resolution cellular atlas represents a community resource and foundation of knowledge to study germ cell development and in vivo gametogenesis.

Title : Single-cell RNA-seq uncovers dynamic processes and critical regulators in mouse spermatogenesis.

Publication date : 09. 2018

Authors : Y Chen, Y Zheng, Y Gao, Z Lin, S Yang, T Wang, Q Wang, N Xie, R Hua, M Liu, J Sha, MD Griswold, J Li, F Tang, MH Tong

A systematic interrogation of male germ cells is key to complete understanding of molecular mechanisms governing spermatogenesis and the development of new strategies for infertility therapies and male contraception. Here we develop an approach to purify all types of homogeneous spermatogenic cells by combining transgenic labeling and synchronization of the cycle of the seminiferous epithelium, and subsequent single-cell RNA-sequencing. We reveal extensive and previously uncharacterized dynamic processes and molecular signatures in gene expression, as well as specific patterns of alternative splicing, and novel regulators for specific stages of male germ cell development. Our transcriptomics analyses led us to discover discriminative markers for isolating round spermatids at specific stages, and different embryo developmental potentials between early and late stage spermatids, providing evidence that maturation of round spermatids impacts on embryo development. This work provides valuable insights into mammalian spermatogenesis, and a comprehensive resource for future studies towards the complete elucidation of gametogenesis.

Title : Transcriptomic analysis of mRNA expression and alternative splicing during mouse sex determination.

Publication date : 12. 2018

Authors : L Zhao, C Wang, ML Lehman, M He, J An, T Svingen, CM Spiller, ET Ng, CC Nelson, P Koopman

Mammalian sex determination hinges on sexually dimorphic transcriptional programs in developing fetal gonads. A comprehensive view of these programs is crucial for understanding the normal development of fetal testes and ovaries and the etiology of human disorders of sex development (DSDs), many of which remain unexplained. Using strand-specific RNA-sequencing, we characterized the mouse fetal gonadal transcriptome from 10.5 to 13.5 days post coitum, a key time window in sex determination and gonad development. Our dataset benefits from a greater sensitivity, accuracy and dynamic range compared to microarray studies, allows global dynamics and sex-specificity of gene expression to be assessed, and provides a window to non-transcriptional events such as alternative splicing. Spliceomic analysis uncovered female-specific regulation of Lef1 splicing, which may contribute to the enhanced WNT signaling activity in XX gonads. We provide a user-friendly visualization tool for the complete transcriptomic and spliceomic dataset as a resource for the field.

Title : Identification of dynamic undifferentiated cell states within the male germline.

Publication date : 07. 2018

Authors : HM La, JA Mäkelä, AL Chan, FJ Rossello, CM Nefzger, JMD Legrand, M De Seram, JM Polo, RM Hobbs

The role of stem cells in tissue maintenance is appreciated and hierarchical models of stem cell self-renewal and differentiation often proposed. Stem cell activity in the male germline is restricted to undifferentiated A-type spermatogonia (A

Title : The transcription factor SOX30 is a key regulator of mouse spermiogenesis.

Publication date : 05. 2018

Authors : D Zhang, D Xie, X Lin, L Ma, J Chen, D Zhang, Y Wang, S Duo, Y Feng, C Zheng, B Jiang, Y Ning, C Han

The postmeiotic development of male germ cells, also known as spermiogenesis, features the coordinated expression of a large number of spermatid-specific genes. However, only a limited number of key transcription factors have been identified and the underlying regulatory mechanisms remain largely unknown. Here, we report that SOX30, the most-divergent member of the Sry-related high-motility group box (SOX) family of transcription factors, is essential for mouse spermiogenesis. The SOX30 protein was predominantly expressed in spermatids, while its transcription was regulated by retinoic acid and by MYBL1 before and during meiosis.

Title : Transcriptome analysis of the adult human Klinefelter testis and cellularity-matched controls reveals disturbed differentiation of Sertoli- and Leydig cells.

Publication date : 05. 2018

Authors : SB Winge, MD Dalgaard, KG Belling, JM Jensen, JE Nielsen, L Aksglaede, MH Schierup, S Brunak, NE Skakkebæk, A Juul, E Rajpert-De Meyts, K Almstrup

The most common human sex chromosomal disorder is Klinefelter syndrome (KS; 47,XXY). Adult patients with KS display a diverse phenotype but are nearly always infertile, due to testicular degeneration at puberty. To identify mechanisms causing the selective destruction of the seminiferous epithelium, we performed RNA-sequencing of 24 fixed paraffin-embedded testicular tissue samples. Analysis of informative transcriptomes revealed 235 differentially expressed transcripts (DETs) in the adult KS testis showing enrichment of long non-coding RNAs, but surprisingly not of X-chromosomal transcripts. Comparison to 46,XY samples with complete spermatogenesis and Sertoli cell-only-syndrome allowed prediction of the cellular origin of 71 of the DETs. DACH2 and FAM9A were validated by immunohistochemistry and found to mark apparently undifferentiated somatic cell populations in the KS testes. Moreover, transcriptomes from fetal, pre-pubertal, and adult KS testes showed a limited overlap, indicating that different mechanisms are likely to operate at each developmental stage. Based on our data, we propose that testicular degeneration in men with KS is a consequence of germ cells loss initiated during early development in combination with disturbed maturation of Sertoli- and Leydig cells.