Function of Nanos2 in the male germ cell lineage in mice

Abstract

Nanos is known as an evolutionarily conserved RNA-binding protein, the function of which is implicated in germ cell development. This includes the maintenance of both the primordial germ cells (PGCs) and germline stem cells. In mice, Nanos2 exhibits a unique feature in which its expression is induced only in the germ cells within the sexually determined male gonad. Nanos2 promotes male germ cell differentiation, while simultaneously suppressing a female program. In addition, Nanos2 is also expressed in the spermatogonial stem cells and functions as an intrinsic factor to maintain the stem cell population during spermatogenesis. Detailed cytological and biochemical analyses in embryonic male gonads in the mouse have revealed that Nanos2 localizes to the P-bodies, a center of RNA processing. It has also been shown that the Nanos2 interacts with protein components of the deadenylation complex involved in the initial step of the RNA degradation pathway.

Fig. 1

Developmental pathways involved in the sexual differentiation of germ cells. Once PGCs are formed, they proliferate and migrate toward the future gonads in which somatic cells commence sexual differentiation by expressing either male or female promoting factors. Retinoic acid (RA) is produced in both male and female gonads, but is destroyed by CYP26b1, which is specifically expressed in the male gonad. RA induces Stra8 expression in female germ cells, which then enter meiosis. In contrast, Stra8 expression is suppressed by RA degradation via Cyp26b1 and later by Nanos2 expression in germ cells. Male germ cells do not enter meiosis during embryogenesis. The expression of Nanos3 and Nanos2 is schematically indicated

Fig. 2

Scheme of the functions of Nanos2 during the sexual differentiation of germ cells. In wild-type male germ cells, Nanos2 promotes male-type differentiation, whereas the absence of Nanos2 in these cells (Nanos2-KO) results in the abnormal induction of meiosis. The forced expression of Nanos2 in female germ cells causes the suppression of meiosis

Fig. 3

Genetic experiments that demonstrate the function of the Nanos2 in spermatogonial stem cells in the adult mouse testes. A The conditional knockout of the Nanos2 gene was induced by TM injection of 4-week-old male mice containing a floxed Nanos2 and ERT2Cre (a). After 4 weeks, almost all of the germ cells were depleted from the seminiferous tubules (b). B The continuous expression of Nanos2 was achieved by crossing CAG-floxed CAT-Nanos2 transgenic and Nanos3-cre mice. Nanos2 expression is maintained continuously in all germ cells of the progeny from the embryonic stages. In the wild-type male testes, the number of Nanos2-positive cells (red) is low (c). However, these cells are markedly increased and occupy the entire tubular surface in the induced testes (d). The forced expression of Nanos2 (green) results in the upregulation of PLZF (magenta), a marker of undifferentiated spermatogonia, including stem cells (e). C A cell lineage tracing experiment conducted by crossing a Nanos2-MCM [MerCreMer, a tamoxifen (TM) inducible Cre] mouse with a Rosa26 reporter mouse. Following the transient administration of TM in the 6-week-old male progeny, β-gal activity was detectable at 7 weeks (f) and 18 weeks (g). The presence of β-gal-positive patches that contained spermatogenic germ cells at all stages of development indicates that Nanos2 is expressed in the stem cell population

Fig. 4

A proposed model for the molecular functions of Nanos2. In male mouse germ cells, RA may reach and activate the transcription of meiotic genes such as Stra8 after E13.5 when Cyp26b1 expression begins to decrease. However, in wild-type mice, these transcripts would be incorporated into the CCR4-NOT deadenylation complex via a Nanos2-mediated mechanism and loaded onto P-bodies where RNAs are degraded by the enzymes contained therein. This results in the suppression of meiosis (X is an unidentified Nanos2-interacting protein) (a). In contrast, RNAs are not loaded onto P-bodies in the absence of Nanos2 and may thus go to the polysome and be translated into the proteins required for meiosis (b)