NANOS2 acts downstream of glial cell line-derived neurotrophic factor signaling to suppress differentiation of spermatogonial stem cells.

Stem cells are maintained by both stem cell-extrinsic niche signals and stem cell-intrinsic factors. During murine spermatogenesis, glial cell line-derived neurotrophic factor (GDNF) signal emanated from Sertoli cells and germ cell-intrinsic factor NANOS2 represent key regulators for the maintenance of spermatogonial stem cells. However, it remains unclear how these factors intersect in stem cells to control their cellular state. Here, we show that GDNF signaling is essential to maintain NANOS2 expression, and overexpression of Nanos2 can alleviate the stem cell loss phenotype caused by the depletion of Gfra1, a receptor for GDNF. By using an inducible Cre-loxP system, we show that NANOS2 expression is downregulated upon the conditional knockout (cKO) of Gfra1, while ectopic expression of Nanos2 in GFRA1-negative spermatogonia does not induce de novo GFRA1 expression. Furthermore, overexpression of Nanos2 in the Gfra1-cKO testes prevents precocious differentiation of the Gfra1-knockout stem cells and partially rescues the stem cell loss phenotypes of Gfra1-deficient mice, indicating that the stem cell differentiation can be suppressed by NANOS2 even in the absence of GDNF signaling. Taken together, we suggest that NANOS2 acts downstream of GDNF signaling to maintain undifferentiated state of spermatogonial stem cells.

Sada A ，Hasegawa K ，Pin PH ，Saga Y ... -  《-》

The RNA-binding protein NANOS2 is required to maintain murine spermatogonial stem cells.

Sada A ，Suzuki A ，Suzuki H ，Saga Y ... -  《-》

GDNF family receptor alpha1 phenotype of spermatogonial stem cells in immature mouse testes.

Buageaw A ，Sukhwani M ，Ben-Yehudah A ，Ehmcke J ，Rawe VY ，Pholpramool C ，Orwig KE ，Schlatt S ... -  《BIOLOGY OF REPRODUCTION》

Distribution of GFRA1-expressing spermatogonia in adult mouse testis.

Grasso M ，Fuso A ，Dovere L ，de Rooij DG ，Stefanini M ，Boitani C ，Vicini E ... -  《-》

Gdnf-Gfra1 pathway is expressed in a spermatogenetic-dependent manner and is regulated by Fsh in a fish testis.

What makes the spermatogonial stem cells (SSCs) self-renew or differentiate to produce spermatozoa is barely understood, in particular in nonmammalian species. Our research explores possible regulations of the SSC niche in teleost, locally by paracrine factors and peripherally by hormonal regulation. In the present study, we focus on the Gdnf-Gfra1 pathway that plays a major role in the regulation of SSC self-renewal in mammals. We describe a complex evolution of the genes encoding for Gdnf and Gfra1 proteins in trout with the emergence of three gdnf and two gfra1 paralogs. Using quantitative PCR measurements in isolated testicular cell populations, the gdnfb paralog was found expressed in A-spermatogonia and probably in another testicular cell type. In contrast, the transcript of gfra1a, the Gdnf receptor, was preferentially expressed in a population of undifferentiated A-spermatogonia (und A-Spg) separated by centrifugal elutriation. These und A-Spg also demonstrated high stemness potential in transplantation studies and preferentially expressed nanos2, a putative SSC marker in trout (Bellaiche et al., Biol Reprod 2014; 90:79). Flow cytometer experiments demonstrate that only a subfraction of und A-Spg express Gfra1. In trout, spermatogenesis develops along a strict annual cycle, and gdnfb and its receptor were expressed in a spermatogenetic activity-dependent manner. In particular, a dramatic increase of the gdnfb transcript coincided with the progressive cessation of rapid spermatogonial proliferation and of meiosis toward the end of the reproductive cycle. Together these results suggest that, in trout, Gdnfb is involved in the repression of und A-Spg differentiation. Fsh is an endocrine regulator of SSCs self-renewal through the up-regulation of Gdnf in rodents. We demonstrate that in trout, in vitro Fsh treatment stimulated the expression of the gfra1a1 but not of its ligand, gdnfb. Fsh treatment also stimulated the proliferation of und A-Spg cocultured with testicular somatic cells. Based on those results, the Gfra1-positive cells could correspond to the putative SSCs in rainbow trout, and we propose that the balance between SSC self-renewal and differentiation during the trout spermatogenetic cycle is under paracrine regulation by Gdnfb, which represses, and under peripheral regulation by Fsh via the control of gfra1a1 expression.

Bellaïche J ，Goupil AS ，Sambroni E ，Lareyre JJ ，Le Gac F ... -  《-》