An S-opsin knock-in mouse (F81Y) reveals a role for the native ligand 11-cis-retinal in cone opsin biosynthesis.
In absence of their natural ligand, 11-cis-retinal, cone opsin G-protein-coupled receptors fail to traffic normally, a condition associated with photoreceptor degeneration and blindness. We created a mouse with a point mutation (F81Y) in cone S-opsin. As expected, cones with this knock-in mutation respond to light with maximal sensitivity red-shifted from 360 to 420 nm, consistent with an altered interaction between the apoprotein and ligand, 11-cis-retinal. However, cones expressing F81Y S-opsin showed an ∼3-fold reduced absolute sensitivity that was associated with a corresponding reduction in S-opsin protein expression. The reduced S-opsin expression did not arise from decreased S-opsin mRNA or cone degeneration, but rather from enhanced endoplasmic reticulum (ER)-associated degradation of the nascent protein. Exogenously increased 11-cis-retinal restored F81Y S-opsin protein expression to normal levels, suggesting that ligand binding in the ER facilitates proper folding. Immunohistochemistry and electron microscopy of normal retinas showed that Mueller cells, which synthesize a precursor of 11-cis-retinal, are closely adjoined to the cone ER, so they could deliver the ligand to the site of opsin synthesis. Together, these results suggest that the binding of 11-cis-retinal in the ER is important for normal folding during cone opsin biosynthesis.
Center for Neuroscience, University of California, Davis, California 95618, USA.
Christine
C
Insinna
Lauren L
LL
Daniele
Jason A
JA
Davis
DeLaine D
DD
Larsen
Colleen
C
Kuemmel
Jinhua
J
Wang
Sergei S
SS
Nikonov
Barry E
BE
Knox
Edward N
EN
Pugh
D000465:Algorithms; D000818:Animals; D030801:Animals, Genetically Modified; D015153:Blotting, Western; D055724:Electrophysiological Phenomena; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D008027:Light; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008854:Microscopy, Electron; D009154:Mutation / Q000502:physiology; D055355:Opsins / Q000096:biosynthesis* / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D017949:Retinal Cone Photoreceptor Cells / Q000378:metabolism*; D017948:Retinal Rod Photoreceptor Cells / Q000378:metabolism; D012172:Retinaldehyde / Q000502:physiology*
D055355:Opsins; D043562:Receptors, G-Protein-Coupled; D012172:Retinaldehyde