The objective of the research is to elucidate mechanisms used for signal transduction and membrane excitability in the nervous system using a combined molecular, biochemical and genetic approach. The results will be useful in understanding abnormalities in the human visual and nervous systems. Our efforts will be focused on the process of phototransduction in the Drosophila eye. The nature of two genetic loci affecting membrane excitability will also be investigated. In order to accomplish our overall objectives we will: (1) Attempt to isolate the genes encoding each of the several different rhodopsins expected to exist in Drosophila. (2) Determine the DNA sequences of the these rhodopsin genes and thereby the amino acid sequences of the proteins they encode. (3) Determine which rhodopsins are expressed in which classes of photoreceptors. (4) Determine the ability of rhodopsins to function in photoreceptor cells in which they are not normally expressed; and if feasible use such ectopic expression to study the spectral sensitivities and other properties of these rhodopsins. (5) Mutate selected amino acids in a rhodopsin protein by site-directed in vitro mutagenesis of its gene and study the properties of the altered protein following re-introduction of the mutant gene into flies by P-element-mediated gene transfer. (6) Characterize in detail the transient receptor potential (trp) gene and its product. (7) Characterize in detail the ninaC gene and its product. (8) Isolate and catalog genes encoding other products involved in the phototransduction process. (9) Characterize some of the genes isolated as described in specific aim (8), choosing those genes which, based on genetic and biochemical criteria, most warrant further work. (10) Isolate and characterize the genes bang-senseless (bss) and easily-shocked (eas). (11) Finally, for each of the genes that we study in detail we will ask whether homologous genes can be identified in vertebrates using methods of nucleic acid sequence cross-homology and antibody cross-reaction.
