The long-term goal of this project is to elucidate the molecular mechanism(s) controlling stem cell proliferation in response to developmental cues. Stem cells are unique precursors that are self-renewing and give rise to various cell types. Insufficient stem cell division leads to birth defects, while uncontrolled division leads to many types of cancer. Thus, stem ceils have an enormous impact on the health, viability and maintenance of all organisms. In Drosophila, subsets of neuronal stem cells, or neuroblasts, are quiescent upon larval hatching and resume cell division at specific developmental stages; the timing of proliferation is affected by mutations in trol. Trol activates division in quiescent neuroblasts. We have shown that trol encodes a homolog of mammalian Perlecan, a co-receptor for growth factors. Our genetic and biochemical analyses suggest that the signaling pathways activated by two growth factors, Bnl (the Drosophila FGF) and Hh, drive initiation of division in the regulated cells. Signaling by these pathways is modulated by Trol. We have begun to produce fusion proteins and generate domain-specific antibodies. Both types of reagents will be used to probe the structure of mutant Trol proteins and the specificity of Trol-FGF and Trol-Hh binding. We will use the antibodies and a trol-GAL4 transgene to follow trol expression. We will examine how each pathway acts in concert or independently to affect division of specific neuroblasts or neuroblast subsets by altering the levels of ligands and other components, characterizing the expression pattern of pathway proteins, and blocking activation of the FGF pathway. To elucidate how signaling specificity is attained, we will assay for allele-specific interactions between trol and bnl or hh, ascertain the effect of over-expressing trol domains in weak ligand mutants, study the consequences of simultaneously changing levels of both Bnl and Hh, and determine the role of sugar moieties in signaling specificity. These studies will provide important insights into the mechanism(s) by which stem cell division is controlled by developmental cues--a process of fundamental importance in all animals, including humans.