Best's macular dystrophy (BMD), a disease with many similarities to age-related macular degeneration (AMD). AMD is the leading cause of blindness in the western world and is currently untreatable. By understanding how mutations in bestrophin cause BMD we will gain insights into the causes of AMD with the expectation that the knowledge gained will lead to effective therapies for this disease. A distinguishing symptom of BMD is a depressed light response in the electro-oculogram without alterations in the electro-retinogram, a condition mimicked by the toxic responses induced by ocular iron overload, and drugs that affect iron metabolism. Thus the studies outlined in this proposal are designed to determine a function for bestrophin guided by the hypothesis that bestrophin is involved in the regulation of iron homeostasis or transcytotic transport by the retinal pigment epithelium (RPE). This hypothesis is reinforced by the presence of several potential iron binding motifs in the amino acid sequence of bestrophin.
Specific aims focus on: The subcellular localization and developmental expression of bestrophin; determination of bestrophin function in vitro; determination of bestrophin function in vivo; and biochemical characterization of bestrophin. The localization of bestrophin will be determined by immunofluorescence microscopy using antibodies raised against bestrophin, or bestrophin fused with green fluorescent protein. The function of bestrophin will be tested in assays of iron binding, ferroxidase/ferrireductase activity, and transport and uptake of iron by the RPE. The effects of mutant bestrophin on the phagocytic pathway of RPE cells, such as delayed kinetics of outer segment degradation and altered pH of endosomal/phagosomal compartments will be tested. BMD associated mutants of bestrophin will be expressed in the RPE of mice and rats using adenovirus mediated gene transfer, followed by histologic, electrophysiologic, and biochemical analysis of the effects on lipofuscin accumulation and retinoid processing.
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