Among cell lineages, germ cells are unique in that they can generate a new organism. In males, germline stem cells provide a source of undifferentiated cells that allow spermatogenesis to proceed throughout the period of sexual maturity. Cells committed to differentiate enter the meiotic pathway, which comprises a unique program of gene expression and chromatin remodeling1. To maintain the stem cell pool, however, some germ cells must remain undifferentiated and proliferate through cyclic mitotic divisions. How does each spermatogonial stem cell decide whether to proliferate or differentiate? The molecular mechanisms controlling this delicate balance are largely unknown. New studies by F. William Buaas and colleagues and José Costoya and colleagues published in this issue2,3 provide clues to the mechanisms required for self-renewal of spermatogonial stem cells by showing that the transcriptional repressor Plzf is required for stem cell maintenance.

An epigenetic connection

Plzf (promyelocytic leukemia zinc-finger) belongs to the POK (POZ and Krüppel) family of transcriptional repressors. In addition to nine Krüppel-type sequence-specific zinc fingers, Plzf contains a conserved POZ (poxvirus and zinc finger) domain in its N terminus. This domain, common to several zinc finger–containing transcription factors, mediates protein-protein interactions and allows POZ domain proteins to participate in various differentiation pathways, including hematopoiesis, adipogenesis, hippocampal neurogenesis, osteoclastogenesis and muscle differentiation.

Earlier studies highlighted the role of Plzf in regulating differentiation. First, Plzf is expressed in early, but not differentiated, hematopoietic cells4, suggesting that Plzf is involved in stem cell maintenance. In addition, Plzf regulates genes involved in cellular proliferation and differentiation (cyclin A2, Myc and Hox genes). Finally, targeted deletion of Plzf in mice disrupts patterning of the limb and axial skeleton5. These features are reminiscent of the luxoid mouse, a mutant described 50 years ago for its limb abnormalities and recessive skeletal phenotype. The studies of Buaas et al. and Costoya et al. now show that luxoid mutants and Plzf-null mice share similar defects in sperm production that are due to an inability of spermatogonial stem cells to self-renew. Consistent with these shared phenotypes, Buaas et al. show that the luxoid mutation results from a frameshift mutation in Zfp145, which encodes Plzf2.

A twist in this story is that Plzf probably influences the epigenetic program of spermatogonial cells. Although direct proof is not given in these studies2,3, previous work showed that the POZ domain of Plzf recruits members of the mammalian Polycomb family, such as BMI1 (refs. 6,7). Polycomb proteins maintain stable and heritable repression of several developmental genes. Recruitment of BMI1 by Plzf results in the subsequent recruitment of histone deacetylases8, thereby linking epigenetic modifications to transcriptional control. Thus, it is reasonable to hypothesize that Plzf-dependent histone deacetylases impose specific chromatin remodeling events that contribute to the decision between differentiation and self-renewal in spermatogonial stem cells.

The reports by Buaas et al. and Costoya et al. highlight the importance of epigenetic modifications and transcriptional regulation as key mechanisms for stem cell maintenance. As several POZ transcriptional regulators function in cellular differentiation, POZ-dependent epigenetic modifications may have a more common role in directing stem cell behavior. This possibility becomes more promising when one considers that the totipotency of germ cells may be epigenetically regulated through DNA methylation.

Signaling and self-renewal

The behavior of germ cells in the seminiferous tubules is largely controlled by the surrounding somatic Sertoli cells. In mice, GDNF (glial cell line–derived neurotrophic factor) produced by Sertoli cells regulates cell fate decisions in undifferentiated spermatogonia9. Studies in fruit flies identified the importance of somatic cell EGF (epidermal growth factor) receptor and Raf activity in determining the differentiation capacity of stem cells10,11. Furthermore, maintenance of the stem cell population in fruit flies seems to be secured by activation of the JAK-STAT pathway in germ cells by the unpaired ligand produced by somatic cells of the testis12. In flies, the translational repressor Nanos is also essential for conserving stem cell status13, and the centrosomal protein centrosomin and the tumor suppressor APC (adenomatous polyposis coli) determine the fate of male germline stem cells by regulating mitotic spindle orientation during asymmetric cell division14.

Despite progress in understanding the mechanisms by which external signals regulate stem cell renewal, little is known about the cell-autonomous factors that control this process in mammalian germ cells. Thus, Plzf is a notable example of a cell-autonomous germ cell factor required for spermatogonial stem cell maintenance. But it remains to be seen how Plzf interacts with known signaling pathways, particularly in response to Sertoli-derived signals such as GDNF and stem cell factor (SCF; Fig. 1).

Figure 1: Some known signaling cascades that control spermatogonial cell fate, and that could converge on Plzf, are schematically represented.
figure 1

Sertoli cells produce SCF, the natural ligand of c-Kit, a tyrosine kinase receptor located on the surface of spermatogonia. Sertoli cells produce SCF when induced by follicle stimulating hormone (FSH), a pituitary hormone released in response to hypothalamic signals. SCF binding to c-Kit elicits dimerization of the receptor and consequent activation of the phosphoinositide 3-kinase (PI3K) pathway, which in turn triggers the Akt-PKB kinase. This cascade directly regulates the apoptosis-survival decision, as Akt-PKB regulates proteins of the Bcl2-Bax family. The SCF-c-Kit pathway is paralleled by the GDNF-Ret system. GDNF is released by Sertoli cells and binds to the oncoprotein Ret, which is also coupled to the PI3K-PKB signaling pathway.

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Stem cells and cancer

The involvement of Plzf in cancer brings additional importance to its identification as a regulator of stem cell differentiation. Chromosomal translocations that fuse Plzf and RARα (retinoid acid receptor α) are associated with acute promyelocytic leukemia15. The ability of Plzf to control stem cell maintenance may be perturbed in acute promyelocytic leukemia, thus providing leukemic progenitors with a proliferative advantage. The Plzf interaction partner BMI1 is required for maintenance of both hematopoietic and leukemic stem cells6,7. Understanding the molecular programs controlling stem cell self-renewal will have an additional impact on development of anticancer therapies.

The unique role of germ cells in transmitting the genome from one generation to the next emphasizes the importance of studying germline stem cells. The multilevel regulatory mechanisms governing germline stem cell self-renewal and differentiation are beginning to be uncovered. The further use of mouse models will provide crucial insights into the genetic and epigenetic components that constitute the complex molecular network underlying stem cell biology.