During hermaphrodite vulval development six vulval precursor cells (VPCs, P3.p through P8.p) are competent to generate vulval tissue and adopt an invariant pattern of three cell fates. The anchor cell secretes the LIN-3 EGF growth factor that activates in P6.p the RTK/RAS/MAPK pathway to specify the primary (1) cell fate. Subsequently, a lateral signal from P6.p induces the secondary (2) cell fate in the neighboring cells P5.p and P7.p via LIN-12 Notch. Several negative regulators of the EGFR/RAS/MAPK signaling pathway, such as DEP-1 and LIP-1 are up-regulated in P5.p and P7.p to inhibit the primary fate specification in these VPCs. Due to genetic redundancy, single mutants of
dep-1(lf) or
lip-1(lf) do not show a defect in vulval development. However,
dep-1(lf);
lip-1(lf) double mutants exhibit an adjacent primary fate (APF) phenotype, in which P5.p and P7.p descendents adopt a hybrid cell fate with mixed 2 and 1 characteristics (Berset et al., 2005). In order to identify new genes that regulate the 1 versus 2 cell fate decision in P5.p and P7.p, two sensitized forward genetic screens were performed in a clonal manner that allowed the isolation of mutations which cause sterility in addition to the vulval phenotype. In a first screen,
dep-1(lf) animals were mutagenized and the progeny of 2040 F1 clones was screened for mutants showing an APF phenotype. A second screen was carried out in a
lip-1(lf) background where the progeny of 2600 F1 clones was screened for the APF phenotype. The expression analysis of
lin-11p::GFP, a molecular marker for the 2 cell fate, contributed to the identification of the three most promising mutants
zh79,
zh83 and
zh85. By using a combination of automated fragment-length polymorphism (FLP) mapping, complementation tests and DNA sequencing, the affected genes of the two mutants
zh83 and
zh85 have been identified. Details about the genes identified in these screens will be presented.