Identification of Stem Cells During Prepubertal Spermatogenesis via Monitoring of Nucleostemin Promoter Activity

Abstract

The nucleostemin (NS) gene encodes a nucleolar protein found at high levels in several types of stem cells and tumor cell lines. The function of NS is unclear but it may play a critical role in S-phase entry by stem/progenitor cells. Here we characterize NS expression in murine male germ cells. Although NS protein was highly expressed in the nucleoli of all primordial germ cells, only a limited number of gonocytes showed NS expression in neonatal testes. In adult testes, NS protein was expressed at high levels in the nucleoli of spermatogonia and primary spermatocytes but at only low levels in round spermatids. To evaluate the properties of cells expressing high levels of NS, we generated transgenic reporter mice expressing green fluorescent protein (GFP) under the control of the NS promoter (NS-GFP Tg mice). In adult NS-GFP Tg testes, GFP and endogenous NS protein expression were correlated in spermatogonia and spermatocytes but GFP was also ectopically expressed in elongated spermatids and sperm. In testes of NS-GFP Tg embryos, neonates, and 10-day-old pups, however, GFP expression closely coincided with endogenous NS expression in developing germ cells. In contrast to a previous report, our results support the existence in neonatal testes of spermatogonial stem cells with long-term repopulating capacity. Furthermore, our data show that NS expression does not correlate with cell-cycle status during prepuberty, and that strong NS expression is essential for the maintenance of germline stem cell proliferation capacity. We conclude that NS is a marker of undifferentiated status in the germ cell lineage during prepubertal spermatogenesis. STEM CELLS 2008; 26:3237-3246 Disclosure of potential conflicts of interest is found at the end of this article.

Neonatal Gonocyte Differentiation in Mongolian GerbilMeriones unguiculatusInvolves Asynchronous Maturation of Seminiferous Cords and Rapid Formation of Transitional Cell Stage

“…This finding corroborates previous studies indicating that prespermatogenic germ cells are a heterogeneous population differing not only in their localization in seminiferous epithelium and morphology but also in their expression of specific molecular markers such as transcription factor OCT-4, tyrosine kinase receptor c-KIT and Melanoma antigen-A4 (MAGE-A4) (Fukuda et al, 1975;Gaskell et al, 2004;Yoshida et al, 2006;Ohmura et al, 2008).…”

Identification of side population cells in mouse primordial germ cells and prenatal testis

“…Ohumura and colleagues recently showed that a nucleolar protein called nucleostemin, found at high levels in several types of stem cells, identifies germ cells with undifferentiated status and long-term testis repopulating capability in prenatal gonads (Ohmura et al, 2008). Interestingly, the expression pattern of this protein closely resembles that of the SP phenotype described here.…”

“…A previous study indicated that, in the adult testis, the SP fraction also contains stem/progenitors for androgen-producing Leydig cells in addition to SSCs (Lo et al, 2004). It would be interesting to test whether embryonic Leydig progenitor cells are also present in the SP fraction from prenatal testes.Ohumura and colleagues recently showed that a nucleolar protein called nucleostemin, found at high levels in several types of stem cells, identifies germ cells with undifferentiated status and long-term testis repopulating capability in prenatal gonads (Ohmura et al, 2008). Interestingly, the expression pattern of this protein closely resembles that of the SP phenotype described here.…”

Turning a new page on nucleostemin and self-renewal

“…Nucleostemin was discovered because of its high expression in neuroepithelial stem or progenitor cells purified from the fetal forebrain (Tsai and McKay, 2002) and was later found to be highly expressed in other types of stem cells, tumors, and tumorinitiating cells or cancer stem cells (CSCs) (Baddoo et al, 2003;Lin et al, 2010;Ohmura et al, 2008;Tamase et al, 2009;Tsai and McKay, 2002;Yamashita et al, 2013). By contrast, in differentiated cells and tissues, this protein is expressed at a much lower level, except for those undergoing regeneration Maki et al, 2007;Shugo et al, 2012;Siddiqi et al, 2008).…”