Characterization of Insect Rpr, Hid, and Grim Homologs

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DESCRIPTION (provided by the applicant): Apoptosis is a vital process in both vertebrates and invertebrates. Functional and evolutionary conservation of apoptotic proteins allows the use of invertebrates as mammalian models to expedite the understanding of cancer biology and embryogenesis. The Drosophila melanogaster pro-death proteins RPR, HID and GRIM maintain their biological functions in both mammalian and invertebrate systems indicating that homologs to these proteins mat exist. To date, no homologs of RPR, HID and GRIM have been identified, even in other insects. Preliminary data from Southern hybridizations of insect genomic DNA shows that at least in other Drosophila subspecies, homologs to these death proteins exist. Initially, homologs from different Drosophila subspecies will be used as probes to screen the invertebrates, Tribolium and Sppodoptera by Southern blot, colony hybridization and degenerate PCR. Homologs will be cloned and sequenced. Protein-protein interactions of the homologs will be assessed using yeast-two hybrid analysis and immunoprecipitation. Finally, functional analysis of the homologs will be addresses by protein over-expression in invertebrat cells to analyze their roles in apoptosis, and RNA interference (RNAi) will evaluate loss of function phenotypes.