No abstract

Mammalian spermatogenesis is maintained by a continuous supply of differentiating cells from self-renewing stem cells. The stem cell activity resides in a small subset of primitive germ cells, the undifferentiated spermatogonia. However, the relationship between the establishment of this population and the initiation of differentiation in the developing testes remains unclear. In this study, we have investigated this issue by using the unique expression of Ngn3, which is expressed specifically in the undifferentiated spermatogonia, but not in the differentiating spermatogonia or their progenitors, the gonocytes. Our lineage analyses demonstrate that the first round of mouse spermatogenesis initiates directly from gonocytes, without passing through the Ngn3-expressing stage (Ngn3 -lineage). By contrast, the subsequent rounds of spermatogenesis are derived from Ngn3-positive undifferentiated spermatogonia, which are also immediate descendents of the gonocytes and represent the stem cell function (Ngn3 + lineage). Thus, in mouse spermatogenesis, the state of the undifferentiated spermatogonia is not an inevitable step but is a developmental option that ensures continuous sperm production. In addition, the segregation of gonocytes into undifferentiated spermatogonia (Ngn3 + lineage) or differentiating spermatogonia (Ngn3 -lineage) is topographically related to the establishment of the seminiferous epithelial cycle, thus suggesting a role of somatic components in the establishment of stem cells.

Interleukin 2 (IL-2), a T cell-derived cytokine, targets a variety of cells to induce their growth, differentiation, and functional activation. IL-2 inserts signals into the cells through IL-2 receptors expressed on cell surfaces to induce such actions. In humans, the functional IL-2 receptor consists of the subunit complexes of the alpha, beta and gamma chains, or the beta and gamma chains. The third component, the gamma chain, of IL-2 receptor plays a pivotal role in formation of the full-fledged IL-2 receptor, together with the beta chain, the gamma chain participates in increasing the IL-2 binding affinity and intracellular signal transduction. Moreover, the cytokine receptors for at least IL-2, IL-4, IL-7, IL-9, and IL-15 utilize the same gamma chain as an essential subunit. Interestingly, mutations of the gamma chain gene cause human X-linked severe combined immunodeficiency (XSCID) characterized by a complete or profound T cell defect. Among the cytokines sharing the gamma chain, at least IL-7 is essentially involved in early T cell development in the mouse organ culture system. The molecular identification of the gamma chain brought a grasp of the structures and functions of the cytokine receptor and an in-depth understanding of the cause of human XSCID. To investigate the mechanism of XSCID and development of gene therapy for XSCID, knockout mice for the gamma chain gene were produced that showed similar but not exactly the same phenotypes as human XSCID.

In mammalian testis, a typical stem cell system ensures continuous spermatozoa production. Lines of experiments have demonstrated that stem cell activity resides in the most primitive small subset of germ cells, that is, A(s) (A(single)), A(pr) (A(paired)), and A(al) (A(aligned)) spermatogonia, also collectively called undifferentiated spermatogonia. However, their cellular or molecular nature is largely to be elucidated because a gene that is specifically expressed in these cells has not yet been identified, which makes it difficult to study them. In this study, we demonstrate that a class B basic helix-loop-helix (bHLH) transcription factor neurogenin3 (ngn3) is expressed specifically in A(s), A(pr), and A(al) spermatogonia because ngn3 is expressed in c-Kit negative spermatogonia throughout the seminiferous cycle, and transgenic labeling with GFP revealed connection of 1, 2, 4, 8, 16, or 32 ngn3-positive cells via intercellular bridges. ngn3 is first expressed at the prepubertal stage in c-Kit negative prespermatogonia. Lineage tracing, using the Cre-loxP system, demonstrates that ngn3-positive germ cells give rise to eventually all the spermatogenesis in mature testis. To our knowledge, ngn3 is the first reported gene that delineates these earliest stages of spermatogenesis. Considering its molecular nature, ngn3 could be involved in their differentiation control. Moreover, visualization with GFP and targeting expression of exogenous genes are valuable tools to investigate the mammalian spermatogenic stem cell system.