Photic generation of 11-cis-retinal in bovine retinal pigment epithelium

Jianye Zhang,Elliot H Choi,Aleksander Tworak,David Salom,Henri Leinonen,Christopher L Sander,Thanh V Hoang,James T Handa,Seth Blackshaw,Grazyna Palczewska,Philip D Kiser,Krzysztof Palczewski

doi:10.1074/jbc.ra119.011169

Abstract

Photoisomerization of the 11-cis-retinal chromophore of rod and cone visual pigments to an all-trans-configuration is the initiating event for vision in vertebrates. The regeneration of 11-cis-retinal, necessary for sustained visual function, is an endergonic process normally conducted by specialized enzyme systems. However, 11-cis-retinal also can be formed through reverse photoisomerization from all-trans-retinal. A nonvisual opsin known as retinal pigment epithelium (RPE)-retinal G-protein-coupled receptor (RGR) was previously shown to mediate visual chromophore regeneration in photic conditions, but conflicting results have cast doubt on its role as a photoisomerase. Here, we describe high-level production of 11-cis-retinal from RPE membranes stimulated by illumination at a narrow band of wavelengths. This activity was associated with RGR and enhanced by cellular retinaldehyde-binding protein (CRALBP), which binds the 11-cis-retinal produced by RGR and prevents its re-isomerization to all-trans-retinal. The activity was recapitulated with cells heterologously expressing RGR and with purified recombinant RGR. Using an RGR variant, K255A, we confirmed that a Schiff base linkage at Lys-255 is critical for substrate binding and isomerization. Single-cell RNA-Seq analysis of the retina and RPE tissue confirmed that RGR is expressed in human and bovine RPE and Müller glia, whereas mouse RGR is expressed in RPE but not in Müller glia. These results provide key insights into the mechanisms of physiological retinoid photoisomerization and suggest a novel mechanism by which RGR, in concert with CRALBP, regenerates the visual chromophore in the RPE under sustained light conditions.

Highlights

Photoisomerization of the 11-cis-retinal chromophore of rod and cone visual pigments to an all-trans-configuration is the initiating event for vision in vertebrates
When retinal pigment epithelium (RPE) microsomes were exposed to white light, we observed the formation of 11-cis-retinal, in amounts that fluctuated from experiment to experiment
Production of 11-cis-retinal was not observed in darkness (Fig. S2, a and b), whereas trace amounts of cis-retinals observed in the samples even before illumination (Fig. S2, c and d) were most likely derived from the incomplete depletion of retinoids in RPE microsomes pre-treated with intense UV light at 302 nm

Summary

Introduction

Photoisomerization of the 11-cis-retinal chromophore of rod and cone visual pigments to an all-trans-configuration is the initiating event for vision in vertebrates. One source of regenerated 11-cis-retinal is a metabolic pathway referred to as the retinoid (visual) cycle (Fig. 1) [4] with a critical step that generates 11-cis-retinol from all-trans-retinyl esters via the action of a retinoid isomerase called RPE65 [5,6,7] This activity is consistent with an earlier proposal that light-independent isomerization of all-trans- to 11-cis-retinoids in the eye occurs at the alcohol oxidation state [8]. The eye has mechanisms to limit retinal illuminance and visual pigment activation, for example by pupillary constriction, bright light exposure can result in high isomerization rates that may exceed the regeneration capacity of the visual cycle, for foveal cones [21] This imbalance may be countered by the photochemical production of 11-cis-retinal under sustained light conditions.

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