The long-term objective of the proposed research is to understand the molecular mechanisms underlying vision in the fruitfly, Drosophila melanogaster. The goal of the current proposal is to understand the functions of a gene, ninaC, which is required for signal transduction in photoreceptor cells (phototransduction) and to prevent retinal degeneration. The gene encodes two highly related proteins, p132 and p174, with linked domains homologous to protein kinases and the head region of myosin heavy chains. The two NINAC isoforms are the major calmodulin. The approach to studying ninaC takes advantage of a combination of molecular, biochemical, genetic, electrophysiological, histological and germline transformation techniques. To study the functions of ninaC we plan to: 1) test the hypothesis that the calmodulin bound to the two different calmodulin binding sites has different functions, 2) identify the rhabdomere localization signal (rhabdomeres are the photoreceptor cell microvillar structures), 3) test the kinase regulates subcellular localization or calmodulin binding, 4) test the proposal that the PEST sequence targets NINAC for degradation, 5) test the hypothesis that the NINAC proteins are bona fide actin-based motors by characterizing the enzymatic properties and testing for in vitro motility, 6) test the hypothesis that pp28 is a target for the NINAC kinase activity, and 7) identify and characterize other Drosophila retinal calmodulin binding proteins. The results of the proposed experiments should contribute not only to understanding visual physiology and the functions of protein-calmodulin complexes, but perhaps also to a more general understanding of the roles of unconventional myosin and proteins kinases. Retinal degeneration is a major health problem; however, in most cases the causes of retinal degeneration are not understood. The degeneration might be due to a requirement for p174 to link the actin filaments in the rhabdomere to the membrane. A number of proteins, such as dystrophia, bridge actin filaments with the plasma membrane and are required to maintain the integrity of the cell. Thus, a description of the mechanism underlying the degeneration of fly photoreceptor cells might also contribute to understanding the bases of diseases which lead to the degeneration of other cell types as well.
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