Enhanced Shutoff of Phototransduction in Transgenic Mice Expressing Palmitoylation-deficient Rhodopsin*♦

Zhongyan Wang,Xiao-Hong Wen,Zsolt Ablonczy,Rosalie K Crouch,Clint L Makino,Janis Lem

doi:10.1074/jbc.m502588200

Abstract

Palmitoylation is a reversible, post-translational modification observed in a number of G-protein-coupled receptors. To gain a better understanding of its role in visual transduction, we produced transgenic knock-in mice that expressed a palmitoylation-deficient rhodopsin (Palm(-/-)). The mutant rhodopsin was expressed at wild-type levels and showed normal cellular localization to rod outer segments, indicating that neither rhodopsin stability nor its intracellular trafficking were compromised. But Palm(-/-) rods had briefer flash responses and reduced sensitivity to flashes and to steps of light. Upon exposure to light, rhodopsin became phosphorylated at a faster rate in mutant than in wild-type retinas. Since quench of rhodopsin begins with its phosphorylation, these results suggest that palmitoylation may modulate rod photoreceptor sensitivity by permitting rhodopsin to remain active for a longer period.

Highlights

Palmitoylation is a reversible, post-translational modification observed in a number of G-protein-coupled receptors
Palmitoylation is a covalent, post-translational modification occurring at the C terminus in many G-protein coupled receptors (GPCR)1 [1]
Abrogation of Rhodopsin Palmitoylation in PalmϪ/Ϫ Mice— Palmitoylation-deficient rhodopsin animals were produced by mutation of the cysteine residues at positions 322 and 323 of rhodopsin to non-palmitoylatable threonine and serine sites, respectively

Summary

Introduction

Palmitoylation is a reversible, post-translational modification observed in a number of G-protein-coupled receptors. In vitro studies of several GPCRs in which the palmitoylated cysteine(s) was mutated have shown changes in G-protein coupling efficiency and selectivity, receptor phosphorylation and desensitization, and cell surface expression and trafficking [1]. Palmitates, bound to cysteines 322 and 323 through a reversible thioester bond [4], insert into the membrane and anchor the C terminus of rhodopsin [5, 6], creating a fourth intracellular domain (Fig. 1) Residues in this domain are important for transducin activation [7, 8]. We examine the role of receptor palmitoylation in vivo using the rod-specific GPCR rhodopsin as the subject of study. Our studies reveal palmitoylation-deficient (PalmϪ/Ϫ) rods to be less sensitive to light, due to increased rhodopsin phosphorylation

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Conclusion