Abstract
The mammalian germ cells, cell assemblies, tissues, and organs during development and maturation have been extensively studied at the tissue level. However, to investigate and understand the fundamental insights at the molecular basis of germ and stem cells, their cell fate plasticity, and determination, it is of most importance to analyze at the large scale on the single-cell level through different biological windows. Here, modern molecular techniques optimized for single-cell analysis, including single fluorescence-activated cell sorting (FACS) and single-cell RNA sequencing (scRNA-seq) or microfluidic high-throughput quantitative real-time polymerase chain reaction (qRT-PCR) for single-cell gene expression and liquid chromatography coupled to tandem mass spectrometry (LC-MSMS) for protein profiling, have been established and are still getting optimized.
This review aims on describing and discussing recent single-cell expression profiling and proteomics of different types of human germ cells, including primordial germ cells (PGCs), spermatogonial stem cells (SSCs), human adult germ stem cells (haGSCs), and oocytes.
Keywords
Abbreviations
- FACS:
-
Fluorescence-activated cell sorting
- haGSC:
-
Human adult germ stem cell
- hESC:
-
Human embryonic stem cells
- hFib:
-
Human fibroblast
- hPSC:
-
Human pluripotent stem cell
- hSSC:
-
Human spermatogonial stem cell
- LC-MSMS:
-
Liquid chromatography mass spectrometry
- MACS:
-
Magnetic activated cell sorting
- MSCs:
-
Mesenchymal stem cells
- hPGC:
-
Human primordial germ cell
- qRT-PCR:
-
Quantitative real-time polymerase chain reaction
- scRNA-seq:
-
Single-cell RNA sequencing
- hSSC:
-
Human spermatogonial stem cell
References
Chikhovskaya, J. V., Jonker, M. J., Meissner, A., Breit, T. M., Repping, S., & van Pelt, A. M. (2012). Human testis-derived embryonic stem cell-like cells are not pluripotent, but possess potential of mesenchymal progenitors. Human Reproduction, 27, 210–221.
Choi, I., Carey, T. S., Wilson, C. A., & Knott, J. G. (2012). Transcription factor AP-2gamma is a core regulator of tight junction biogenesis and cavity formation during mouse early embryogenesis. Development, 139, 4623–4632.
Conrad, S., Azizi, H., Hatami, M., Kubista, M., Bonin, M., Hennenlotter, J., Sievert, K. D., & Skutella, T. (2016). Expression of genes related to germ cell lineage and Pluripotency in single cells and colonies of human adult germ stem cells. Stem Cells International, 2016, 8582526.
Gerovska, D., & Arauzo-Bravo, M. J. (2016). Does mouse embryo primordial germ cell activation start before implantation as suggested by single-cell transcriptomics dynamics? Molecular Human Reproduction, 22, 208–225.
Giritharan, G., Li, M. W., Di Sebastiano, F., Esteban, F. J., Horcajadas, J. A., Lloyd, K. C., Donjacour, A., Maltepe, E., & Rinaudo, P. F. (2010). Effect of ICSI on gene expression and development of mouse preimplantation embryos. Human Reproduction, 25, 3012–3024.
Gkountela, S., Li, Z., Vincent, J. J., Zhang, K. X., Chen, A., Pellegrini, M., & Clark, A. T. (2013). The ontogeny of cKIT+ human primordial germ cells proves to be a resource for human germ line reprogramming, imprint erasure and in vitro differentiation. Nature Cell Biology, 15, 113–122.
Gonzalez, R., Griparic, L., Vargas, V., Burgee, K., Santacruz, P., Anderson, R., Schiewe, M., Silva, F., & Patel, A. (2009). A putative mesenchymal stem cells population isolated from adult human testes. Biochemical and Biophysical Research Communications, 385, 570–575.
Graf, T., & Enver, T. (2009). Forcing cells to change lineages. Nature, 462, 587–594.
Grassetti, D., Paoli, D., Gallo, M., D’Ambrosio, A., Lombardo, F., Lenzi, A., & Gandini, L. (2012). Protamine-1 and -2 polymorphisms and gene expression in male infertility: An Italian study. Journal of Endocrinological Investigation, 35, 882–888.
Grindberg, R. V., Yee-Greenbaum, J. L., McConnell, M. J., Novotny, M., O’Shaughnessy, A. L., Lambert, G. M., Arauzo-Bravo, M. J., Lee, J., Fishman, M., Robbins, G. E., et al. (2013). RNA-sequencing from single nuclei. Proceedings of the National Academy of Sciences of the United States of America, 110, 19802–19807.
Guo, F., Yan, L., Guo, H., Li, L., Hu, B., Zhao, Y., Yong, J., Hu, Y., Wang, X., Wei, Y., et al. (2015). The Transcriptome and DNA Methylome landscapes of human primordial germ cells. Cell, 161, 1437–1452.
Hayashi, K., Ohta, H., Kurimoto, K., Aramaki, S., & Saitou, M. (2011). Reconstitution of the mouse germ cell specification pathway in culture by pluripotent stem cells. Cell, 146, 519–532.
Hermann, B. P., Mutoji, K. N., Velte, E. K., Ko, D., Oatley, J. M., Geyer, C. B., & McCarrey, J. R. (2015). Transcriptional and translational heterogeneity among neonatal mouse spermatogonia. Biology of Reproduction, 92, 54.
Hough, S. R., Thornton, M., Mason, E., Mar, J. C., Wells, C. A., & Pera, M. F. (2014). Single-cell gene expression profiles define self-renewing, pluripotent, and lineage primed states of human pluripotent stem cells. Stem Cell Reports, 2, 881–895.
Huang, S., Ernberg, I., & Kauffman, S. (2009). Cancer attractors: A systems view of tumors from a gene network dynamics and developmental perspective. Seminars in Cell & Developmental Biology, 20, 869–876.
Huang, L., Ma, F., Chapman, A., Lu, S., & Xie, X. S. (2015). Single-cell whole-genome amplification and sequencing: Methodology and applications. Annual Review of Genomics and Human Genetics, 16, 79–102.
Hughes, C. S., Foehr, S., Garfield, D. A., Furlong, E. E., Steinmetz, L. M., & Krijgsveld, J. (2014). Ultrasensitive proteome analysis using paramagnetic bead technology. Molecular systems biology 10, 757.
Jang, S., Choubey, S., Furchtgott, L., Zou, L.N., Doyle, A., Menon, V., Loew, E.B., Krostag, A.R., Martinez, R.A., Madisen, L., et al. (2017). Dynamics of embryonic stem cell differentiation inferred from single-cell transcriptomics show a series of transitions through discrete cell states. eLife, 6.
Kooistra, S. M., Thummer, R. P., & Eggen, B. J. (2009). Characterization of human UTF1, a chromatin-associated protein with repressor activity expressed in pluripotent cells. Stem Cell Research, 2, 211–218.
Kristensen, D. M., Nielsen, J. E., Skakkebaek, N. E., Graem, N., Jacobsen, G. K., Rajpert-De Meyts, E., & Leffers, H. (2008). Presumed pluripotency markers UTF-1 and REX-1 are expressed in human adult testes and germ cell neoplasms. Human Reproduction, 23, 775–782.
Li, L., Dong, J., Yan, L., Yong, J., Liu, X., Hu, Y., Fan, X., Wu, X., Guo, H., Wang, X., et al. (2017). Single-cell RNA-Seq analysis maps development of human Germline cells and gonadal niche interactions. Cell Stem Cell, 20, 891–892.
Liu, Q., Li, Y., Feng, Y., Liu, C., Ma, J., Li, Y., Xiang, H., Ji, Y., Cao, Y., Tong, X., et al. (2016). Single-cell analysis of differences in transcriptomic profiles of oocytes and cumulus cells at GV, MI, MII stages from PCOS patients. Scientific Reports, 6, 39638.
Luetjens, C. M., Xu, E. Y., Rejo Pera, R. A., Kamischke, A., Nieschlag, E., & Gromoll, J. (2004). Association of meiotic arrest with lack of BOULE protein expression in infertile men. The Journal of Clinical Endocrinology and Metabolism, 89, 1926–1933.
Maekawa, M., Yamamoto, T., Kohno, M., Takeichi, M., & Nishida, E. (2007). Requirement for ERK MAP kinase in mouse preimplantation development. Development, 134, 2751–2759.
Magnusdottir, E., Dietmann, S., Murakami, K., Gunesdogan, U., Tang, F., Bao, S., Diamanti, E., Lao, K., Gottgens, B., & Azim Surani, M. (2013). A tripartite transcription factor network regulates primordial germ cell specification in mice. Nature Cell Biology, 15, 905–915.
Mizrak, S. C., Chikhovskaya, J. V., Sadri-Ardekani, H., van Daalen, S., Korver, C. M., Hovingh, S. E., Roepers-Gajadien, H. L., Raya, A., Fluiter, K., de Reijke, T. M., et al. (2010). Embryonic stem cell-like cells derived from adult human testis. Human Reproduction, 25, 158–167.
Nakamura, T., Yabuta, Y., Okamoto, I., Aramaki, S., Yokobayashi, S., Kurimoto, K., Sekiguchi, K., Nakagawa, M., Yamamoto, T., & Saitou, M. (2015). SC3-seq: A method for highly parallel and quantitative measurement of single-cell gene expression. Nucleic Acids Research, 43, e60.
Neuhaus, N., Yoon, J., Terwort, N., Kliesch, S., Seggewiss, J., Huge, A., Voss, R., Schlatt, S., Grindberg, R. V., & Scholer, H. R. (2017). Single-cell gene expression analysis reveals diversity among human spermatogonia. Molecular Human Reproduction, 23, 79–90.
Novershtern, N., Subramanian, A., Lawton, L. N., Mak, R. H., Haining, W. N., McConkey, M. E., Habib, N., Yosef, N., Chang, C. Y., Shay, T., et al. (2011). Densely interconnected transcriptional circuits control cell states in human hematopoiesis. Cell, 144, 296–309.
Ohinata, Y., Ohta, H., Shigeta, M., Yamanaka, K., Wakayama, T., & Saitou, M. (2009). A signaling principle for the specification of the germ cell lineage in mice. Cell, 137, 571–584.
Okuda, A., Fukushima, A., Nishimoto, M., Orimo, A., Yamagishi, T., Nabeshima, Y., Kuro-o, M., Nabeshima, Y., Boon, K., Keaveney, M., et al. (1998). UTF1, a novel transcriptional coactivator expressed in pluripotent embryonic stem cells and extra-embryonic cells. The EMBO Journal, 17, 2019–2032.
Ooi, S. K., & Bestor, T. H. (2008). The colorful history of active DNA demethylation. Cell, 133, 1145–1148.
Perrett, R. M., Turnpenny, L., Eckert, J. J., O’Shea, M., Sonne, S. B., Cameron, I. T., Wilson, D. I., Rajpert-De Meyts, E., & Hanley, N. A. (2008). The early human germ cell lineage does not express SOX2 during in vivo development or upon in vitro culture. Biology of Reproduction, 78, 852–858.
Robinson, L. L., Gaskell, T. L., Saunders, P. T., & Anderson, R. A. (2001). Germ cell specific expression of c-kit in the human fetal gonad. Molecular Human Reproduction, 7, 845–852.
Saliba, A. E., Westermann, A. J., Gorski, S. A., & Vogel, J. (2014). Single-cell RNA-seq: Advances and future challenges. Nucleic Acids Research, 42, 8845–8860.
Shapiro, E., Biezuner, T., & Linnarsson, S. (2013). Single-cell sequencing-based technologies will revolutionize whole-organism science. Nature Reviews Genetics, 14, 618–630.
Tanay, A., & Regev, A. (2017). Scaling single-cell genomics from phenomenology to mechanism. Nature, 541, 331–338.
Tang, F., Lao, K., & Surani, M. A. (2011). Development and applications of single-cell transcriptome analysis. Nature Methods, 8, S6–11.
Valli, H., Sukhwani, M., Dovey, S. L., Peters, K. A., Donohue, J., Castro, C. A., Chu, T., Marshall, G. R., & Orwig, K. E. (2014). Fluorescence- and magnetic-activated cell sorting strategies to isolate and enrich human spermatogonial stem cells. Fertility and Sterility, 102(566–580), e567.
Virant-Klun, I., Knez, K., Tomazevic, T., & Skutella, T. (2013). Gene expression profiling of human oocytes developed and matured in vivo or in vitro. BioMed Research International, 2013, 879489.
Virant-Klun, I., Leicht, S., Hughes, C., & Krijgsveld, J. (2016). Identification of maturation-specific proteins by single-cell proteomics of human oocytes. Molecular & Cellular Proteomics: MCP, 15, 2616–2627.
von Kopylow, K., Kirchhoff, C., Jezek, D., Schulze, W., Feig, C., Primig, M., Steinkraus, V., & Spiess, A. N. (2010). Screening for biomarkers of spermatogonia within the human testis: A whole genome approach. Human Reproduction, 25, 1104–1112.
von Kopylow, K., Staege, H., Schulze, W., Will, H., & Kirchhoff, C. (2012a). Fibroblast growth factor receptor 3 is highly expressed in rarely dividing human type a spermatogonia. Histochemistry and Cell Biology, 138, 759–772.
von Kopylow, K., Staege, H., Spiess, A. N., Schulze, W., Will, H., Primig, M., & Kirchhoff, C. (2012b). Differential marker protein expression specifies rarefaction zone-containing human Adark spermatogonia. Reproduction, 143, 45–57.
von Kopylow, K., Schulze, W., Salzbrunn, A., & Spiess, A. N. (2016). Isolation and gene expression analysis of single potential human spermatogonial stem cells. Molecular Human Reproduction, 22, 229–239.
Wagner, A., Regev, A., & Yosef, N. (2016). Revealing the vectors of cellular identity with single-cell genomics. Nature Biotechnology, 34, 1145–1160.
Woodworth, M. B., Girskis, K. M., & Walsh, C. A. (2017). Building a lineage from single cells: Genetic techniques for cell lineage tracking. Nature Reviews Genetics, 18, 230–244.
Yosef, N., & Regev, A. (2011). Impulse control: Temporal dynamics in gene transcription. Cell, 144, 886–896.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2017 Springer International Publishing AG
About this paper
Cite this paper
Conrad, S., Azizi, H., Skutella, T. (2017). Single-Cell Expression Profiling and Proteomics of Primordial Germ Cells, Spermatogonial Stem Cells, Adult Germ Stem Cells, and Oocytes. In: Van Pham, P. (eds) Stem Cells: Biology and Engineering. Advances in Experimental Medicine and Biology(), vol 1083. Springer, Cham. https://doi.org/10.1007/5584_2017_117
Download citation
DOI: https://doi.org/10.1007/5584_2017_117
Published:
Publisher Name: Springer, Cham
Print ISBN: 978-3-319-77481-7
Online ISBN: 978-3-319-77482-4
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)