May Berenbaum, University of Illinois at Urbana-Champaign Understanding how plant-feeding insects such as the Lepidoptera, the butterflies and moths, recognize and utilize the plants on which they feed is key to understanding the diversification of this exceptionally species-rich group of organisms. The inability to identify specific genes underlying how hostplants are identified and processed has been a major obstacle; although genome projects provide enormous inventories the challenge remains to assign function to genes affecting behavior and physiology and substantial knowledge gaps characterize the ecology of most model organisms such as Drosophila melanogaster, the "fruit fly" of genetics fame. Key genes mediating plant-insect interactions are the cytochrome P450 monooxygenases, enzymes responsible not only for detoxifying insecticidal plant compounds consumed by leaf-feeding caterpillars but also breaking down plant odorants detected by the antennae and legs of female butterflies searching for plants on which to lay eggs . An extraordinary opportunity for characterizing P450s within a single species that influence hostplant use by caterpillars and hostplant recognition by female butterflies is presented by the black swallowtail Papilio polyxenes. Its chemical ecology is uniquely well-characterized, stimulants for egg-laying as well as toxic chemicals that affect caterpillar growth are known. As well, for a non-model species, its molecular biology is unusually well-documented, with P450 enzymes CYP6B1 and CYP6B3 in caterpillar guts contributing to detoxification of insecticidal compounds in hostplants, and CYP4G28 characterized from the legs of female butterflies possibly involved in hostplant recognition by female butterflies. Thus, the objectives of this SGER are to determine CYP4G28 function by 1. determining whether this gene is expressed specifically in the sensory structures used by female butterflies for "tasting" and recognizing their hostplants prior to laying eggs (using the techniques of immunolocalization and in situ hybridization); 2. molecular modeling to determine whether chemicals known to stimulate egg-laying "fit" into the active site of the CYP4G28 protein; 3. conducting behavioral bioassays to determine if inhibiting the P450s in butterfly forelegs prohibits female butterflies from recognizing their hostplants. Broader impacts resulting from the proposed activities include interdisciplinary training encompassing elements of phytochemistry, molecular biology, and entomology; students will receive broad-based training essential for competitiveness in the 21st century. Findings will be published in scientific journals, in a textbook integrating molecular biology and chemical ecology, and in popular writings and public lectures. Among other potential benefits to society at large will be a greater understanding of the process by which cytochome P450s evolve. These enzymes contribute to the development of insecticide resistance and the acquisition of new hostplants, including crops, phenomena of tremendous ecological and economic importance.
