MOLECULAR ANALYSIS OF OCULAR LENS BEADED FILAMENTS

University Of California At Davis, Davis CA

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Abstract

DESCRIPTION (provided by applicant): We seek to understand the function of Intermediate Filaments in the ocular lens, with a special emphasis on the Beaded Filament, a derivative of the Intermediate Filament which is unique to the lens fiber cell. To accomplish this we propose studies which will: 1. Identify Beaded Filament- and Intermediate Filament Associated Proteins. Identification of such proteins will yield clues to the function of these filaments in the lens, and lead to an understanding of how these filaments integrate into lens cell biology. 2. Identify structural relationships between Beaded/Intermediate Filaments and the lens cell. Identification of structural relationships will again suggest hypotheses of function, and delineate how the filaments are integrated into lens cell biology. 3. Define the molecular relationships of beaded filament proteins in the intact beaded filament, so that the impact of cataract-causing mutations can be understood. 4. Define the dynamics of Beaded/Intermediate Filaments in living lens cells, to gain an understanding of what functions these structures play in the lens. These studies will utilize a range of cell biological and molecular techniques, and take advantage of opportunities provided by knockout mice lacking CP49, CP95, and vimentin. We will employ light and electron microscopy, immunochemical approaches, molecular cloning, protein purification and identification, site-directed spin labeling/electron paramagnetic resonance, confocal microscopy, transfection, and transgenic approaches. Completion of these studies will explain the function of theses proteins/structures in normal lens biology, permitting us to understand how inherited human mutations in these proteins result in cataract.