The expression profile for KIT in male germ cells (present in prospermatogonial precursors, suppressed in undifferentiated spermatogonia and subsequently induced upon transition to a differentiating state) suggests silencing by small RNAs. Mounting evidence indicates small non-coding RNAs, such as microRNAs

INTRODUCTION Male fertility requires continual spermatogenesis, a process dependent on activities of an undifferentiated spermatogonial population composed of spermatogonial stem cells (SSCs) and transient amplifying progenitor spermatogonia (de Rooij and Russell, 2000; Oatley and Brinster, 2012). Self-renewal of SSCs maintains a constant pool from which progenitors will arise and amplify in number before committing to a pathway of terminal differentiation. During steady-state conditions, spermatogenesis initiates when undifferentiated progenitor spermatogonia transition to a differentiating state, thereby committing to eventual formation of spermatozoa. In testes of mice, the timeframe of spermatogenesis is 35 days and constant cycling of the germ cell lineage relies on occurrence of the undifferentiated-todifferentiating spermatogonial transition at a defined interval (Oakberg, 1956; Clermont and Trott, 1969). To provide robustness, a majority of undifferentiated spermatogonia make the transition and only a small subpopulation remains in an undifferentiated state. At present, molecular mechanisms regulating transition from an undifferentiated to a differentiating state in mammalian spermatogonia are undefined. In testes of mammals, a somatic cell population composed of Sertoli, Leydig and myoid cells supports germ cell activities. Signaling from a combination of growth factors secreted from these somatic cells, including glial cell line-derived neurotrophic factor (GDNF), fibroblast growth factor 2 (FGF2), and colony stimulating factor 1 (CSF-1), promotes maintenance and proliferation of the undifferentiated spermatogonial population, including self-renewal of SSCs (Meng et al., 2000; Kubota et al., 2004b; Oatley et al., 2009). In addition, signaling from retinoic acid (RA) induces spermatogonial transition from an undifferentiated to a differentiating state (Morales and Griswold, 1987; van Pelt and de Rooij, 1990b; Van Pelt and De Rooij, 1990a). Thus, undifferentiated spermatogonia are in an active state of mitosis while being primed to initiate differentiation upon activation of RA signaling. A hallmark of the transition to a differentiating state in mammalian spermatogonia is the attainment of KIT receptor expression (Yoshinaga et al., 1991; de Rooij, 1998; SchransStassen et al., 1999). Expression of KIT protein is absent in undifferentiated spermatogonia and becomes first detectable on the surface of differentiating spermatogonia and persists until the leptotene stage of spermatocyte development (Manova et al., 1990; Yoshinaga et al., 1991; Schrans-Stassen et al., 1999). In addition, expression of KIT on the surface of spermatogonia is inversely related to SSC capacity (Shinohara et al., 2000; Kubota et al., 2003). Furthermore, during neonatal testis development the undifferentiated spermatogonial population develops from transition of prospermatogonia (e.g. gonocytes) that are KIT+ (Yoshinaga et al., 1991). Thus, establishment of an undifferentiated state in spermatogonia coincides with suppression of KIT expression, which is maintained until activation of RA signaling induces transition to a differentiating state. The expression profile for KIT in male germ cells (present in prospermatogonial precursors, suppressed in undifferentiated spermatogonia and subsequently induced upon transition to a differentiating state) suggests silencing by small RNAs. Mounting evidence indicates small non-coding RNAs, such as microRNAs 1School of Molecular Biosciences, Center for Reproductive Biology, College of Veterinary Medicine, Washington State University, Pullman, WA 99164, USA. 2Center for Reproductive Biology and Health, Department of Animal Sciences, Pennsylvania State University, University Park, PA 16802, USA.