The distal ends of both male and hermaphrodite germlines are populated by proliferating stem cells that initiate meiotic development as they move proximally, away from the distal tip cell (DTC). We are interested in how the distal germ cells exit from a mitotic proliferative state to meiosis and differentiation. Previous work has shown that a GLP-1/Notch signaling pathway, with the LAG-2 ligand expressed by the DTC and the GLP-1 receptor expressed by the germ cells, functions to promote proliferation (Austin and Kimble, 1987; Lambie and Kimble, 1991). We have shown genetically that
gld-1, encoding a KH domain containing RNA binding protein, promotes entry into meiosis or inhibits proliferation (Francis et al., 1995). Kadyk and Kimble (1998) further demonstrated that
gld-1functions redundantly with
gld-2 for this role such that loss of the activities of both genes results in a germ line tumor of proliferating cells in males and hermaphrodites, similar to that seen when
glp-1 is constitutively activated (Berry et al., 1997). Kadyk and Kimble (1998) also demonstrated that the
gld-2(0)
gld-1(0) tumor is epistatic to
glp-1(0), suggesting that GLP-1/Notch signaling inhibits the activities of
gld-1 and
gld-2. GLD-1 levels are low in the distal end but increase proximally until reaching maximum levels around 21 cell diameters from the DTC, approximately where germ cells appear to initiate meiotic development.
gld-1(
oz10gf) single mutants, which have increased GLD-1 accumulation in the distal end, have a smaller proliferative zone, and
gld-1(
oz10) enhances the weak
glp-1(
bn18) allele, both supporting that GLD-1 spatial regulation is important in regulating the entry into meiosis decision. Crittenden et al. (2002) have also suggested that GLD-1 levels in the distal end are important for regulating entry into meiosis based on analysis of FBF regulation of GLD-1 accumulation. Three lines of evidence suggest that the GLP-1/Notch signaling pathway is involved in inhibiting GLD-1 accumulation in the distal end. First, in the absence of GLP-1 activity GLD-1 levels are increased in the distal end. The lack of GLP-1 activity normally results in a very small germline, therefore we looked at GLD-1 accumulation in
gld-2(0)
gld-1(
q361) mutants with and without GLP-1 activity (in the
gld-1(
q361) allele protein is made but is non functional). In
gld-2(0)
gld-1(
q361) animals GLD-1 protein levels are low in the distal end but increase gradually (same as wild-type), however in
gld-2(0)
gld-1(
q361);
glp-1(0) animals, GLD-1 accumulation is dramatically increased in the distal end. Second, the rise in GLD-1 level is delayed (more proximal) in animals with increased GLP-1 activity in the distal end. Third, GLD-1 levels are low or absent in animals with tumorous germlines due to constitutively activated GLP-1. In order to identify other genes that regulate the entry into meiosis decision, possibly through regulating GLD-1 levels, we conducted a genetic screen looking for mutations that are synthetic tumorous with
gld-2(0). From this, we identified five loss-of-function mutations in a Drosophila Nanos homologue,
nos-3, which was previously identified for its ability to bind FBF (Kraemer et al., 1999). While GLD-1 levels are roughly equivalent to wild-type in single
nos-3(0) and
gld-2(0) mutants, they are dramatically reduced in
gld-2(0);
nos-3(0) double mutants suggesting that
gld-2 and
nos-3 function redundantly to promote GLD-1 accumulation. Therefore
gld-2 and
nos-3 function in opposition to Glp-1/Notch signaling in regulating GLD-1 accumulation in the distal germline.