Lack of Light-driven RGR Interferes with Regeneration of Rhodopsin in Darkness

Abstract

Abstract: : Purpose:RGR (retinal G protein-coupled receptor) is a pigmentepithelial photo-isomerase that converts all-trans retinoids into the 11-cis form. RGR was shown to be essential for the maintainance of 11-cis retinal and rhodopsin steady state levels during exposure of the retina to light (Chen et al., 2001, Nature Genetics, 28, pp. 256-260). We tested whether absence of RGR might also influence rhodopsin regeneration in darkness, which depends on the ‘classical’ visual cycle. Methods:RGR-/- and RGR+/+ mice were exposed to 5000lux of white light for 10 minutes. Rhodopsin was measured after different post-exposure periods in darkness. Retinal function was assessed under the same conditions by ERG. RGR and other proteins of the visual cycle were analysed by Western blotting. Rhodopsin regeneration kinetics in RGR-/- was also compared to those in mice expressing the same variant (Met 450) and amount of RPE65. Results:In RGR deficient mice recovery of rhodopsin in darkness from a strong bleach proceeds about 3 times slower as compared to RGR+/+ mice. This slowed darkadaptation causes a slowed recovery of rod function from bleaching, but leaves cone function virtually unchanged. RGR deficient mice have reduced amounts of RPE65 protein. However, comparison with mice expressing the same amount and variant of RPE65 almost completely excludes reduced RPE65 as reason for the slowed darkadaptation. Other proteins of the visual cycle (IRBP, 11-cis RoDH and CRALBP) are expressed in similar amounts in RGR+/+ and -/- mice. Conclusions: Because RGR is a photoisomerase it should not influence regeneration of rhodopsin in darkness. However, RGR-/- mice show a strongly affected darkadaptation, pointing to an interaction of the 'classical' visual cycle and RGR. Although a good candidate, reduced RPE65 levels in RGR-/- animals seem to contribute only marginally to this phenotype. Analysis of other proteins of the visual cycle and retinoid metabolites is currently in progress.

Keywords: retinal pigment epithelium • retinoids/retinoid binding proteins • dark/light adaptation