Hostname: page-component-78c5997874-fbnjt Total loading time: 0 Render date: 2024-11-10T09:01:39.512Z Has data issue: false hasContentIssue false
  • English
  • Français

Expression and intracellular localization of Nanos2-homologue protein in primordial germ cells and spermatogonial stem cells

Published online by Cambridge University Press:  19 March 2019

Vivek Pandey ,
Anima Tripathi Open the ORCID record for Anima Tripathi [Opens in a new window]  and
Pawan K. Dubey Open the ORCID record for Pawan K. Dubey [Opens in a new window]
Vivek Pandey
Affiliation:
Centre for Genetic Disorders, Institute of Science, Banaras Hindu University, Varanasi-221005, India
Anima Tripathi
Affiliation:
Department of Zoology, MMV-BHU, Varanasi-221005, India
Pawan K. Dubey*
Affiliation:
Centre for Genetic Disorders, Institute of Science, Banaras Hindu University, Varanasi-221005, India
*
Address for correspondence: Pawan K. Dubey. Centre for Genetic Disorders, Institute of Science, Banaras Hindu University, Varanasi-221005, India. Tel: +919451890938. E-mail: pkdubey@bhu.ac.in; pawandubey1981@gmail.com
Get access Rights & Permissions [Opens in a new window]

Summary

The decision by germ cells to differentiate and undergo either oogenesis or spermatogenesis takes place during embryonic development and Nanos plays an important role in this process. The present study was designed to investigate the expression patterns in rat of Nanos2-homologue protein in primordial germ cells (PGCs) over different embryonic developmental days as well as in spermatogonial stem cells (SSCs). Embryos from three different embryonic days (E8.5, E10.5, E11.5) and SSCs were isolated and used to detect Nanos2-homologue protein using immunocytochemistry, western blotting, reverse transcription polymerase chain reaction (RT-PCR) and flow cytometry. Interestingly, Nanos2 expression was detected in PGCs at day E11.5 onwards and up to colonization of PGCs in the genital ridge of fetal gonads. No Nanos2 expression was found in PGCs during early embryonic days (E8.5 and 10.5). Furthermore, immunohistochemical and immunofluorescence data revealed that Nanos2 expression was restricted within a subpopulation of undifferentiated spermatogonia (As, single type A SSCs and Apr, paired type A SSCs). The same results were confirmed by our western blot and RT-PCR data, as Nanos2 protein and transcripts were detected only in PGCs from day E11.5 and in undifferentiated spermatogonia (As and Apr). Furthermore, Nanos2-positive cells were also immunodetected and sorted using flow cytometry from the THY1-positive SSCs population, and this strengthened the idea that these cells are stem cells. Our findings suggested that stage-specific expression of Nanos2 occurred on different embryonic developmental days, while during the postnatal period Nanos2 expression is restricted to As and Apr SSCs.

Keywords

Germ cellsImmunocytochemistryNANOS2RatSpermatogonial stem cells
Type
Research Article
Information
Zygote , Volume 27 , Issue 2 , April 2019 , pp. 82 - 88
Copyright
© Cambridge University Press 2019 

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Asaoka-Taguchi, M, Yamada, M, Nakamura, A, Hanyu, K and Kobayashi, S (1999) Maternal Pumilio acts together with Nanos in germline development in Drosophila embryos. Nat Cell Biol 1, 431–437.Google Scholar
Beer, RL and Draper, BW (2013) Nanos3 maintains germline stem cells and expression of the conserved germline stem cell gene Nanos2 in the zebrafish ovary. Dev Biol 374, 308–318.Google Scholar
Bowles, J, Knight, D, Smith, C, Wilhelm, D, Richman, J, Mamiya, S, Yahiro, K, Chawengsaksophak, K, Wilson, MJ, Rossant, J, Hamada, H and Koopman, P (2006) Retinoid signaling determines germ cell fate in mice. Science 312, 596−600.Google Scholar
Forbes, A and Lehmann, R (1998) Nanos and Pumilio have critical roles in the development and function of Drosophila germline stem cells. Development 125, 679–690.Google Scholar
Haraguchi, S, Tsuda, M, Kitajima, S, Sasaoka, Y, Nomura-Kitabayashid, A, Kurokawa, K and Saga, Y (2003) Nanos1: a mouse Nanos gene expressed in the central nervous system is dispensable for normal development. Mech Dev 120, 721–731.Google Scholar
Julaton, VTA and Pera, RAR (2011) Nanos3 function in human germ cell development. Hum Mol Genet 20, 2238–2250.Google Scholar
Kato, Y, Katsuki, T, Kokubo, H, Masuda, A and Saga, Y (2016) Dazl is a target RNA suppressed by mammalian NANOS2 in sexually differentiating male germ cells. Nat Commun 7, 11272.Google Scholar
Kraemer, B, Critenden, S, Gallegos, M, Moulder, G, Barstead, R, Kimble, J and Wickens, M (1999) NANOS-3 and FBF proteins physically interact to control the sperm-oocyte switch in Caenorhabditis elegans . Curr Biol 9, 1009–1118.Google Scholar
Kusz-Zamelczyk, K, Sajek, M, Spik, A Glazar, R, Jedrzecjczak, P, Latos-Bielenska, A, Kotecki, M, Pawelczyk, L and Jaruzelsaka, J (2013) Mutations of NANOS1, a human homologue of the Drosophila morphogen, are associated with a lack of germ cells in testes or severe oligo-astheno-teratozoospermia. J Med Genet 50, 187–193.Google Scholar
Lai, F, Singh, A and King, ML (2012) Xenopus Nanos1 is required to prevent endoderm gene expression and apoptosis in primordial germ cells. Development 139, 1476–1486.Google Scholar
Liu, W, Li, SZ, Li, Z, Wang, Y, Li, XY, Zhong, JX, Zhang, XJ, Zhang, J, Zhou, L and Gui, JF (2015) Complete depletion of primordial germ cells in an all-female fish leads to sex-biased gene expression alteration and sterile all-male occurrence. BMC Genomics 16, 971.Google Scholar
Mochizuki, K, Sano, H, Kobayashi, S, Nishimiya-Fujisawa, C and Fujisawa, T (2000) Expression and evolutionary conservation of Nanos-related genes in Hydra . Dev Genes Evol 210, 591–602.Google Scholar
Ohta, H, Wakayama, T and Nishimune, Y (2004) Commitment of fetal male germ cells to spermatogonial stem cells during mouse embryonic development. Biol Reprod 70, 1286–1291.Google Scholar
Oulhen, N and Gary, MW (2016) Differential Nanos 2 protein stability results in selective germ cell accumulation in the sea urchin. Dev Biol, 418, 146–156.Google Scholar
Pellegrini, M, Filipponi, D, Gori, M, Barrios, F, Lolicato, F, Grimaldi, P, Rossi, P, Jannini, EA, Geremia, R and Dolci, S (2008) ATRA and KL promote differentiation toward the meiotic program of male germ cells. Cell Cycle 7, 3878–3888.Google Scholar
Peng, JX, Xie, JL, Zhou, L, Hong, YH and Gui, JF (2009) Evolutionary conservation of dazl genomic organization and its continuous and dynamic distribution throughout germline development in gynogenetic gibel carp. J Exp Zool 312, 855–871.Google Scholar
Saba, R, Kato, Y and Saga, Y (2014) NANOS2 promotes male germ cell development independent of meiosis suppression. Dev Biol 385, 32−40.Google Scholar
Sada, A, Suzuki, A, Suzuki, H and Saga, Y (2009) The RNA-binding protein NANOS2 is required to maintain murine spermatogonial stem cells. Science 325, 1394–1398.Google Scholar
Subramaniam, K and Seydoux, G (1999) nos-1 and nos-2, two genes related to Drosophila Nanos, regulate primordial germ cell development and survival in Caenorhabditis elegans . Development 126, 4861–4871.Google Scholar
Suzuki, A and Saga, Y (2008) Nanos2 suppresses meiosis and promotes male germ cell differentiation. Genes Dev 22, 430–435.Google Scholar
Suzuki, A, Tsuda, M and Saga, Y (2007) Functional redundancy among Nanos proteins and a distinct role of Nanos2 during male germ cell development. Development 134, 77–83.Google Scholar
Suzuki, H, Saba, R, Sada, A and Saga, Y (2010) The Nanos3-3′UTR is required for germ cell specific NANOS3 expression in mouse embryos. PLoS One 5, 2.Google Scholar
Tsuda, M, Sasaoka, Y, Kiso, M, Abe, K, Haraguchi, S, Kobayashi, S and Saga, Y (2003) Conserved role of NANOS proteins in germ cell development. Science 301, 1239.Google Scholar
Xu, H, Gui, J and Hong, Y (2005) Differential expression of vasa RNA and protein during spermatogenesis and oogenesis in the gibel carp (Carassius auratus gibelio), a bisexually and gynogenetically reproducing vertebrate. Dev Dyn 233, 872–882.Google Scholar