The research proposed addresses the role of the oligosaccharide chains of rhodopsin in disc membrane morphogenesis. It is proposed that these oligosaccharides participate in adhesional interactions between the closely apposed surfaces of the evaginating plasmalemma which ultimately form the lumenal surfaces of closed discs. The hypothesis will be tested by (1) preventing the attachment of oligosaccharides to opsin by biosynthetic modification of the peptide attachment site, (2) perturbing the normal structure of the attached oligosaccharides by selective inhibition of the oligosaccharide processing enzymes, (3) competing against opsin's oligosaccharides with exogenously supplied carbohydrates of known structure, and (4) removing opsin's oligosaccharides from the plasmalemma surface enzymatically. The effect of each of these treatments on retinas in short-term in vivo cultures will be assessed in comparison with controls by monitoring the incorporation of radiolabeled membrane glycoprotein precursors into newly forming rod outer segment membranes, using biochemical techniques as well as light and electron microscopic autoradiography. The morphology and amount of the newly assembled membranes will be analyzed by electron microscopy. Elucidation of these finer details of the basic mechanisms of disc morphogenesis will provide a more complete understanding of normal photoreceptor cell biology. This may provide clues for discovering heretofore unappreciated potential loci of genetic and metabolic defects which promote the selective dysgenesis or degeneration of photoreceptors in certain human retinal dystrophies.
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