9421740 Simcox A signal transduction pathway involving intracellular Ras is highly conserved and used by a number of receptor-tyrosine kinases including the Drosophila EGF receptor (DER). The long term goal of this work is to understand how signal transduction mediated by ubiquitous DER results in a tissuespecific response. DER has multiple roles in development and is involved in cell survival, differentiation and proliferation. Tissue-specific signaling via the receptor is likely to involve localized aomponents, including ligands. The candidate ligand defective dorsal discs (ddd) is the focus of this proposal. The ddd activity is required for ventral patterning in the embryo and growth of the imaginal wing and haltere discs; ddd transcripts are spatially localized in these tissues. The genetic characterization of ddd shows it functions nonautonomously and interacts with DER. DER is epistatic to ddd which suggests ddd acts upstream of the receptor. Both the nonautonomy and the data also support this idea. The predicted ddd protein has an EGF-like domain, which is characteristic of EGF receptor ligands. Ddd also has an Iglike domain. The neuregulins share this combination of domains. These ligands bind vertebrate Neu, an EGF-receptor relative. Interestingly, DER is equally similar to Neu and the EGF receptor. Thus, the genetic and molecular evidence suggest Ddd is a DER ligand. Here it is proposed to extend the molecular-genetic characterization of ddd and test the hypothesis that Ddd is a DER ligand. The objectives are: (1) To determine the phenotypic effects of expression of ddd transgenes. Constructs based on ddd cDNAs will be used to rescue ddd mutants and to determine the developmental consequence of ectopic ddd expression. (2) To describe the tissue and cell distribution of Ddd throughout development. Polyclonal antisera directed against a Ddd-fusion protein will be used in antibody staining in animals of all developmental stages. The temporal and spatial expression dom ain and subcellular or extracellular location of Ddd will be determined. Ddd expression in other disc mutants will be examined. (3) To test the hypothesis that Ddd is a DER ligand using cell-culture experiments. The ability of Ddd and DER to interact molecularly will be tested in ligandbinding and immunoprecipitation assays. The ability of Ddd to activate DER will be tested by monitoring receptor autophosphorylation. The highly conserved EGF receptor pathway regulates cell multiplication and differentiation. Not surprisingly, genes in the pathway have oncogenic forms. Thus this research on ddd, a candidate ligand for the Drosophila EGF receptor, which is related to both Neu and the EGF receptor, will contribute to the understanding of abnormal, as well as normal, growth control and pattern formation.
