The relationship between opsin overexpression and photoreceptor degeneration

PURPOSE. TO characterize the process by which overexpression of normal opsi n leads to photoreceptor degeneration. METHODS. Three transgenic mouse lines were generated that express different levers of an opsin with three amino acid modifications at the C terminus. These modifications created an epitopic site that can be readily distinguis hed from the endogenous protein using a bovine opsin-specific antibody. Evi dence of degeneration associated with opsin overexpression was provided by anatomic studies and electroretinogram (ERG) recordings. Western blot analy sis was used to con the production of the transgenic opsin, and an enzyme-l inked immunosorbent assay (ELISA) was used to determine the amounts of opsi n overexpressed in each line. Immunocytochemistry was used to determine the cellular localization of transgenic opsin. Amounts of Il-cis retinal were determined by extraction and high-performance liquid chromatography (HPLC). RESULTS. Opsin expression levels in the three lines were found to be 123%, 169%, and 222% of the level measured in nontransgenic animals, providing di rect correlation between the level of transgene expression and the severity of the degenerative phenotype. In the lower expressing lines, ERG a-wave a mplitudes were reduced to less than approximately 30% and 15% of normal val ues, whereas responses of the highest expressing line were indistinguishabl e from noise. In the lowest expressor, a 26% elevation in 11-cis retinal wa s observed, whereas in the medium and the high expressors, 11-cis retinal l evels were increased by only 30% to 33%, well below the 69% and 122% increa ses in opsin levels. CONCLUSIONS. The overexpression of normal opsin induces photoreceptor degen eration that is similar to that seen in many mouse models of retinitis pigm entosa. This degeneration can be induced by opsin levels that exceed by onl y approximately 23% that of the normal mouse retina. Opsin overexpression h as potential implications in retinitis pigmentosa.