Mechanisms for Light-driven Chromophore Synthesis by Müller Cells to Regenerate Cone Opsin and Maintain Cone Sensitivity

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Project Summary / Abstract The goal of this project is to identify three proteins of the intrinsic retinal visual cycle that maintain the sensitivity of cones under daylight conditions. During the previous funding period, the PI found that co- expression of RGR opsin and retinol dehydrogenase-10 (Rdh10) confers upon cells the capacity to take up all-trans-retinol (atROL) from the medium and release 11-cis-retinol (11cROL), only on exposure to visible light. The same atROL to 11cROL conversion activity has been observed by Müller cells, although not previously reported as light dependent. Cones, but not rods, possess an 11cROL- dehydrogenase (11cRDH) activity that allows them to utilize the 11cROL released by Müller cells to make 11cRAL chromophore and thereby regenerate photobleached cone opsins. The long-term goal of this project is to understand the processes that enable continuous cone vision during daylight exposure in mammals. Given the importance of cones to human vision, addressing this deficit in our knowledge is certainly within the NEI mission. Specific Aim 1 is to determine the roles of RGR opsin and Rdh10 in the intrinsic retinal visual cycle. This will be accomplished in a collaborative study using biochemical and physiological methodologies. The central hypothesis for Aim 1 is that RGR opsin and Rdh10 comprise the 11cROL-generator in Müller cells, and that formation of 11cROL by these proteins is driven by light. Specific Aim 2 is to identify the 11cRDH that allows cones to regenerate visual pigment from 11cROL. This will be accomplished using a cDNA expression-screen to test members of the short-chain dehydrogenase/reductase (SDR) family that are expressed in cones. The expected outcome of this study is the identification of three proteins that together comprise the intrinsic retinal visual cycle. These results amount to the uncovering of a new metabolic pathway in the retina. They will greatly extend our knowledge about the regeneration of cone visual pigment in daylight. Also, they will open the door to further genetic and functional biochemical studies into the retinal visual cycle, including the potential to identify new susceptibility loci for inherited retinopathies and maculopathies. Mutations in the human RGR gene have already been associated with retinitis pigmentosa in a small subset of cases. While it has been long appreciated that invertebrates, such as insects, use light to regenerate their visual pigments, this has never been shown for vertebrates. The PI's finding that light drives regeneration of cone visual pigment is therefore a novel and important discovery.