Specific genetic defects are responsible for a large percentage of the retinal diseases that afflict human populations. The long range goal of this research program is to characterize analogous genetically inherited retinal degeneration syndromes in the invertebrate, Drosophila, to better understand the molecular basis of degenerative retinal diseases. During previous grant periods, we determined the molecular basis of four inherited retinal degeneration syndromes in Drosophila. This proposal is focused on the cellular mechanisms by which mutations in one of these genes, rhodopsin, causes retinal disease. Rhodopsin mutations are known to cause retinal degeneration in humans, other vertebrates, and in Drosophila. In most known cases, including the Drosophila mutants we will study, the mutant rhodopsin acts in a dominant fashion to trigger degeneration. Also, the Drosophila mutants share the property with the majority of human mutant genes that they appear to disrupt the posttranslational maturation pathway. The experiments proposed here will investigate the cellular mechanisms and additional gene products involved in rhodopsin maturation and recycling. The project is organized into four specific aims: (1) To examine the dominant nature of certain Drosophila rhodopsin mutants. We will use histological markers to examine the fate of the mutant protein in photoreceptors, determine the effect of the mutant rhodopsin on other membrane proteins, and develop a genetic screen for additional genes required for rhodopsin maturation (2) To identify additional molecular components involved in rhodopsin maturation. Gene products required for membrane protein maturation through the endoplasmic reticulum/Golgi complex and for post-Golgi transport of rhodopsin will be characterized. We will analyze mechanisms involved in rhodopsin transport to the photosensitive membranes and retinal degeneration B protein transport to the specialized endoplasmic reticulum membranes of the photoreceptor. (3) To analyze the enzymatic regulation and substrate specificity of the retinal degeneration C (rdgC) gene. Rhodopsin is thought to be a substrate of the rdgC phosphatase. We will characterize the enzymatic activity of the rdgC protein and analyze mutants of rdgC exhibiting altered regulation. (4) To characterize two gene products required rhodopsin maturation already identified by mutation. These genes are required to generate high levels of rhodopsin, probably by promoting efficient utilization of vitamin A. We will determine if these ones encode proteins that act in photoreceptors, and if so, determine the molecular nature of the encoded gene.
