Identifying genes important for spermatogonial stem cell self-renewal and survival

Abstract

Spermatogonial stem cells (SSCs) are the foundation for spermatogenesis and, thus, preservation of a species. Because of stem cell rarity, studying their self-renewal is greatly facilitated by in vitro culture of enriched biologically active cell populations. A recently developed culture method identified glial cell line-derived neurotrophic factor (GDNF) as the essential growth factor that supports in vitro self-renewal of SSCs and results in an increase in their number. This system is a good model to study mechanisms of stem cell self-renewal because of the well defined culture conditions, enriched cell population, and available transplantation assay. By withdrawing and replacing GDNF in culture medium, we identified regulated expression of many genes by using microarray analysis. The expression levels of six of these genes were dramatically decreased by GDNF withdrawal and increased by GDNF replacement. To demonstrate the biological significance of the identified GDNF-regulated genes, we examined the importance of the most responsive of the six, bcl6b, a transcriptional repressor. By using siRNA to reduce transcript levels, Bcl6b was shown to be crucial for SSC maintenance in vitro. Moreover, evaluation of Bcl6b-null male testes revealed degeneration and͞or absence of active spermatogenesis in 24 ؎ 7% of seminiferous tubules. These data suggest that Bcl6b is an important molecule in SSC self-renewal and validate the biological relevance of the GDNF-regulated genes identified through microarray analysis. In addition, comparison of data generated in this study to other stem cell types suggests that self-renewal in SSCs is regulated by distinctly different molecular mechanisms.glial cell line-derived neurotrophic factor ͉ germ-line stem cell ͉ microarray ͉ mouse

Transcriptome analysis of differentiating spermatogonia stimulated with kit ligand

“…The induction of Plzf, a known marker of spermatogonial stem cells (Buaas et al, 2004;Costoya et al, 2004), is in apparent conflict with the above described pro-differentiative pattern of gene expression set up by KL treatment of spermatogonia. Indeed, Plzf is expressed in gonocytes, and in undifferentiated spermatogonia, but it is thought not to be expressed in kit positive spermatogonia in the adult testis (Oatley et al, 2006). However, Plzf is co-expressed with kit in a subset of spermatogonial stem cells during the first wave of spermatogenesis (Naughton et al, 2006).…”

Xeno-free culture of human spermatogonial stem cells supported by human embryonic stem cell-derived fibroblast-like cells

“…Mammalian SSC culture experiments have proved that cultured SSCs require a feeder layer, but the specific roles of feeder layer cells in the initial and prolonged culture of stem cells are not yet clear. Feeder layer cells provide an ideal environment for the growth of stem cells, including detoxification of the culture medium, the secretion of many unique proteins that participate in cell growth and extracellular matrix formation and remodeling [26]. There are several potential advantages of using hdFs as feeder layer cells to support the growth of hESCs [7], and the growth of human SSCs with hdFs as the feeder layer may prevent the interspecies transfer of viruses, but these advantages are rarely studied in human SSC culture.…”

GDNF Signaling Regulates YBX1/mRNA Interactions in Mouse Spermatogonia.

“…Furthermore, many groups have used microarray analysis to identify SSC transcripts in vivo and in SSC culture [121][122][123][124]. However, the focus of the current study is on what happens to mRNAs in SSCs after they are produced by the transcriptional machinery and before they are translated into functional proteins.…”