The first step in vision is the absorption of light by the chromophore of visual pigments. Vitamin A (retinol) is the precursor for visual pigment chromophore and also has important roles in retina development. Vitamin A is delivered from liver, the main storage site, to the eye by retinol binding protein (RBP), a specialized vitamin A carrier protein in the blood. RBP effectively solubilizes vitamin A, a hydrophobic molecule, and protects it from enzymatic and oxidative damage. In humans, loss of RBP function impairs dark adaptation, and causes progressive atrophy of the retinal pigment epithelium (RPE) in the eye at young ages. Under conditions of vitamin A limitation at which wild-type mice behave normally, RBP knockout mice have rapid vision loss after merely a week of vitamin A deficiency. The RPE is the main cell type in the eye for vitamin A absorption and storage for vision. Overwhelming evidence from many independent laboratories over the past 30 years suggests the existence of a specific RBP receptor that mediates vitamin A uptake in the RPE. Study of the molecular mechanism of vitamin A uptake into RPE has been greatly hampered by the lack of knowledge about the identity of the RBP receptor, which turned out to be extremely difficult to purify or clone. We have identified the RBP receptor using a novel biochemical strategy followed by mass spectrometry and have demonstrated, using a variety of techniques, that it has the expected properties of the RBP receptor. The RBP receptor is a multitransmembrane-domain protein of previously unknown function and is not homologous to any protein of known function. It binds to RBP with high affinity and has robust vitamin A activity from the vitamin A/RBP complex. Unlike most other small molecule delivery systems in eukaryotic cells that depend on extracellular carrier proteins, vitamin A uptake by the RBP receptor does not depend on endocytosis of the carrier protein RBP. We have immunolocalized the RBP receptor to the basolateral membrane of the RPE, a location consistent with its role in interacting with RBP in the choroidal blood for vitamin A uptake. In this proposal, we will elucidate the molecular mechanism of this novel membrane transport system by systematically studying the interaction of RBP with the RBP receptor, the mechanism of vitamin A uptake by the RBP receptor and the intracellular mechanism for the RBP receptor-mediated cellular uptake of vitamin A. Since vitamin A is essential for vision, the human eye is highly sensitive to vitamin A deficiency or to the loss of RBP function. Insufficient vitamin A uptake by the eye can lead to defective dark adaptation, RPE atrophy and even severe blindness. Understanding the molecular mechanism of vitamin A uptake by the eye will have a significant impact on efforts to preserve human vision.
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