Scotopic vision is initiated upon capture of a photon of light by rhodopsin molecules present in rod photoreceptor cells. The activation of the light receptor rhodopsin sets into motion a series of biochemical reactions called phototransduction, which leads to the hyperpolarization of the cell. The long-term goal of this research program is to understand the molecular mechanisms underlying the biochemical events in phototransduction under normal and diseased states. The starting point will be structure-function studies of rhodopsin. The importance of this molecule extends beyond its central role in phototransduction. The rhodopsin gene is a hot spot for mutations causing inherited vision disorders and these mutations are the leading cause of autosomal dominant retinitis pigmentosa, a heterogeneous group of inherited retinal degenerative diseases. Despite the wealth of knowledge available for rhodopsin, an accurate mechanism of its action is still unavailable and the mechanism underlying mutations in the light receptor causing vision disorders is unclear. Our immediate goal is to explore emerging ideas about the system that expand on classical dogma;namely, the notion of multiple active states of rhodopsin and the organization of rhodopsin into clusters of dimers.
The aims of the proposal are thematically linked around understanding the fundamental molecular principles governing the activity of rhodopsin in normal and diseased conditions in people. In the first aim, we will test the implicit assumption made in most studies that the structure and function of human rhodopsin is similar to that of the receptor from better-studied mammalian species (bovine and mouse) used to understand human disease pathology. In the second aim, we will test the hypothesis that there are multiple active states of the receptor and that at least one of these states leads to constitutive activity in a rhodopsn mutant causing congenital stationary night blindness. In the third aim, we will test a putative rhodopsin dimer model and determine whether receptor oligomerization contributes to the phenotype of a rhodopsin mutant causing autosomal dominant retinitis pigmentosa. Significant technological advances are required to overcome the intrinsic difficulties in studying membrane proteins to observe native structural and molecular details that are important to understand the system. Our proposal utilizes several high-resolution biophysical methods including atomic force microscopy, single-molecule force spectroscopy and Forster resonance energy transfer. The combination of these methods with more traditional biochemical, biophysical, and genetic approaches will overcome the limitations of traditional assays alone and allow us to directly test emerging paradigms about rhodopsin structure and function. The successful testing of these new concepts will lead to a more accurate molecular framework to understand the function of the system under normal conditions and dysfunctions in inherited human disease.

Public Health Relevance

We are in an era of research where technology is giving us access to the smallest details of biological systems. Our proposal takes advantage of these technological advances in order to understand the initial events in vision under normal and diseased states. Our studies will reveal fundamental insights about the visual system and will reveal novel strategies to combat retinal disease.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY021731-03
Application #
8528609
Study Section
Special Emphasis Panel (ZRG1-BDPE-J (09))
Program Officer
Neuhold, Lisa
Project Start
2011-09-30
Project End
2016-08-31
Budget Start
2013-09-01
Budget End
2014-08-31
Support Year
3
Fiscal Year
2013
Total Cost
$372,875
Indirect Cost
$135,375
Name
Case Western Reserve University
Department
Ophthalmology
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Whited, Allison M; Park, Paul S-H (2014) Atomic force microscopy: a multifaceted tool to study membrane proteins and their interactions with ligands. Biochim Biophys Acta 1838:56-68
Park, Paul Shin-Hyun (2014) Constitutively active rhodopsin and retinal disease. Adv Pharmacol 70:1-36
Kawamura, Shiho; Gerstung, Moritz; Colozo, Alejandro T et al. (2013) Kinetic, energetic, and mechanical differences between dark-state rhodopsin and opsin. Structure 21:426-37
Kawamura, Shiho; Colozo, Alejandro T; Ge, Lin et al. (2012) Structural, energetic, and mechanical perturbations in rhodopsin mutant that causes congenital stationary night blindness. J Biol Chem 287:21826-35
Park, P S-H (2012) Ensemble of G protein-coupled receptor active states. Curr Med Chem 19:1146-54