SPRY4-dependent ERK negative feedback demarcates functional adult stem cells in the male mouse germline.

Overview

Niche-derived growth factors (GFs) support self-renewal of mouse spermatogonial stem and progenitor cells (SSCs) through ERK MAPK signaling and other pathways. At the same time, dysregulated GF-dependent signaling has been associated with loss of stem cell activity and aberrant differentiation. We hypothesized that GF signaling through the ERK MAPK pathway in SSCs is tightly regulated within a narrow range through distinct intracellular negative feedback regulators (NFRs). Evaluation of candidate ERK MAPK-responsive genes known to dampen downstream signaling revealed robust induction of specific NFRs, including Spry4, in cultured mouse SSCs in response to GDNF or FGF2. Undifferentiated spermatogonia in vivo exhibited high levels of Spry4 mRNA. Quantitative single cell analysis of ERK MAPK signaling in SSC cultures revealed both dynamic signaling patterns in response to GFs and disruption of such effects when Spry4 was ablated, due to dysregulation of ERK MAPK downstream of RAS. Whereas NFR expression decreased during differentiation, loss of Spry4 shifted cell fate toward early differentiation with concomitant loss of stem cell activity. Finally, a mouse Spry4 reporter line revealed that the adult SSC population in vivo is demarcated by strong Spry4 promoter activity. Collectively, our data suggest that negative feedback-dependent regulation of ERK MAPK is critical for preservation of SSC fate within the mammalian testis.