Dimerization of visual pigments in vivo

Abstract

It is a deeply engrained notion that the visual pigment rhodopsin signals light as a monomer, even though many G protein-coupled receptors are now known to exist and function as dimers. Nonetheless, recent studies (albeit all in vitro) have suggested that rhodopsin and its chromophore-free apoprotein, R-opsin, may indeed exist as a homodimer in rod disk membranes. Given the overwhelmingly strong historical context, the crucial remaining question, therefore, is whether pigment dimerization truly exists naturally and what function this dimerization may serve. We addressed this question in vivo with a unique mouse line (S-opsin + Lrat −/− ) expressing, transgenically, short-wavelength-sensitive cone opsin (S-opsin) in rods and also lacking chromophore to exploit the fact that cone opsins, but not R-opsin, require chromophore for proper folding and trafficking to the photoreceptor's outer segment. In R-opsin's absence, S-opsin in these transgenic rods without chromophore was mislocalized; in R-opsin's presence, however, S-opsin trafficked normally to the rod outer segment and produced functional S-pigment upon subsequent chromophore restoration. Introducing a competing R-opsin transmembrane helix H1 or helix H8 peptide, but not helix H4 or helix H5 peptide, into these transgenic rods caused mislocalization of R-opsin and S-opsin to the perinuclear endoplasmic reticulum. Importantly, a similar peptidecompetition effect was observed even in WT rods. Our work provides convincing evidence for visual pigment dimerization in vivo under physiological conditions and for its role in pigment maturation and targeting. Our work raises new questions regarding a potential mechanistic role of dimerization in rhodopsin signaling.rhodopsin | cone opsin | dimerization | protein trafficking R hodopsin and cone pigments mediate scotopic and photopic vision, respectively. They consist of opsin, the apo-protein, and 11-cis-retinal, a vitamin A-based chromophore. Light absorption by 11-cis-retinal triggers a conformational change in opsin, which in turn initiates a G protein-coupled receptor (GPCR) signaling pathway to lead to vision. Indeed, rhodopsin signaling is a prominent prototypical GPCR pathway from which a huge quantity of mechanistic details about such signaling in general has emerged. All along, it is a dogma that rhodopsin exists and functions as a monomer (1-6). About a decade ago, evidence began to emerge that rhodopsin may exist as a dimer, based on atomic force microscopy and cross-linking experiments performed on rod outersegment (ROS) disk membranes (7-9). However, this concept remains highly controversial because of the lack of in vivo evidence and also is puzzling because, unlike many GPCRs, monomeric rhodopsin is fully functional with respect to coupling to G protein (2, 4-6, 10) and to interactions with rhodopsin kinase and arrestin (11,12). In vivo evidence, albeit of paramount importance, is also challenging, because rhodopsin always exists as a single isoform in rod photoreceptors, thus making homomeric, higher-or...

The human non-visual opsin OPN3 regulates pigmentation of epidermal melanocytes through interaction with MC1R

“…Members of the opsin family, in particular, have been recently shown to form hetero-and homomeric complexes [39,40]. To determine whether OPN3 and MC1R colocalize to the same cellular compartments, we expressed tagged OPN3 and MC1R in HeLa cells.…”

“…This raises the intriguing question of whether OPN3 functions in the brain similar to how it does in melanocytes: could OPN3 interact and negatively regulate the function of other melanocortin receptors like MC3R? OPN3 is not the only opsin that functions as part of a dimeric complex; a functional homomeric complex has also been reported for rhodopsin (OPN2) and cone opsin (OPN1) [40]. These complexes are mediated by residues within the fifth transmembrane domain of human red and green opsins [39], and by transmembrane domain one and helix eight for rhodopsin dimerization [40].…”

“…OPN3 is not the only opsin that functions as part of a dimeric complex; a functional homomeric complex has also been reported for rhodopsin (OPN2) and cone opsin (OPN1) [40]. These complexes are mediated by residues within the fifth transmembrane domain of human red and green opsins [39], and by transmembrane domain one and helix eight for rhodopsin dimerization [40]. Whether OPN3 and MC1R directly interact through domain coupling or are part of a larger complex was beyond the scope of our study, but will be determined by future studies.…”

Human nonvisual opsin 3 regulates pigmentation of epidermal melanocytes through functional interaction with melanocortin 1 receptor

“…Several recent reports indicate that GPCRs can form functional and physical interactions (37,38). Members of the opsin family, in particular, have recently been shown to form heteroand homomeric complexes (39,40). To determine whether OPN3 and MC1R colocalize to the same cellular compartments, we expressed epitope-tagged OPN3 and MC1R in HeLa cells.…”

A G Protein-Coupled Receptor Dimerization Interface in Human Cone Opsins

“…A recent study of murine S-opsin reported cotrafficking with R-opsin, suggesting that it dimerizes with rhodopsin in a mouse model. 48 Another study noted blue cone opsin aggregation due to a Phe-rich region in this receptor, 49 but the aggregated protein was located in the ER and trans-Golgi network instead of the plasma membrane. Using the approach outlined above, we found that red cone opsin was significantly dimerized, and that the average cross-correlation was higher than that of rhodopsin.…”