% Department of Biology, York University, 4700 Keele St. Ontario, Canada, # Institute of Histology and Embryology, Faculty of Medicine, University of Lisbon, Ave. Prof. Egas Moniz, 1699 Lisboa Codex, Portugal, @ Department of Genetics andMolecular Biology, Guelph University, Guelph, Ontario, Canada, § Federal Institute for Neurobiology, Department of Neurochemistry and Molecular Biology, Brenneckestr. 6, Postfach 1860, D-39008 Magdeburg, Germany,
There are two naturally occurring phenotypic variants in foraging behaviour. One variant is the 'rover', another variants is the 'sitter'. The larval rovers travel significantly further than the sitters while foraging on a yeast and water paste. Genetic and molecular studies demonstrated that the major foraging gene is dg2, which is a putative serine/theronine cGMP-dependent protein kinase (PkG). Chaser (Csr) is a second gene identified which effects foraging behaviour. Csr-1, -2, and -3 alleles were isolated by a gamma mutagenesis screen for a change in behaviour from sitter to rover. The lethal-tagging technique was used to revert Csr-3 through a mutagensis screen for sitter behaviour. Deficiency and complementation analysis demonstrated that the mutations in the reverted Csr-3 line are in the 95F-96A1 region. Three Drosophila genes, als, da2 and sbd, have been cloned and mapped to the 96A region by in-situ hybridization. All of the 3 genes code for nicotinic acetycholine receptor subunits (AChR). We used these genes to probe Csr and Csr revertant RNA on northern. We found an overexpression of the als 10.5kb transcript in Csr-3. The effects of the expression of the als on larval path-length was also investigated using the UAS-GAL4 system. Three independent transgenic lines (UAS-als/da2GAL4) showed that the overexpression of als causes Csr-like behaviour results in a significant increase in travel path-length. This strongly suggestes that the Csr-3 is a mutation in an alpha like nAChR. We are interesting how DG2 and ALS effect foraging behaviour. It is known that the phosphorylation of the AChR can increase the efficiency of assembly of different subunits into the AChR complex and it can also accelerate its rate of desensitization. Hence, it would be interesting to know whether or not DG2 directly interacts and phosphorylate ALS or any of the subunits of the the AChR. The cytoplasmic loop between the M3 and M4 in the ALS and ARD subunits of the nAChR harbor potential phosphorylation sites for protein kinase A (PkA) and PkG. We used the two hybrid system to investigate these potential interactions by constructing dg2 into a PAS2 vector and the cDNA fragments between the M3 and M4 of the als and ard into a pGAD424 and a pACT2 vector respectively. The constructs were cotransformed into the yeast strain CG1945 to test for possible interactions. No interaction was detected between the DG2 and ALS, or DG2 and ARD, indicating that other proteins are likely to mediate the signal transduction between the DG2 and ALS to effect the foraging behaviour. To identify these proteins, we used dg2-PAS2 and als-pGBT9 to screen a two hybrid system library. We have isolated 59 putative positive clones to be further analyzed.