Modulating expression of peripherin/rds in transgenic mice: critical levels and the effect of overexpression.

PURPOSE Mutations in the photoreceptor-specific protein peripherin/rds are associated with multiple retinal diseases. To date, attempts to achieve complete structural and functional rescue in animal models of peripherin/rds-induced retinal degeneration have not been successful. Gene therapy-directed approaches have been hindered by the haploinsufficiency phenotype, which dictates well-regulated expression of peripherin/rds protein levels. METHODS Using a transgenic mouse line expressing wild-type peripherin/rds (NMP), the authors evaluated the critical in vivo level of peripherin/rds needed to maintain photoreceptor structure and ERG function and assessed the consequences of peripherin/rds overexpression in both rods and cones by Western blot and immunoprecipitation analyses, immunohistochemistry, electron microscopy, and electroretinography. The NMP transgene included a C-terminal modification (P341Q) to facilitate detection of the transgenic protein in the presence of wild-type peripherin/rds, using the monoclonal antibody 3B6. RESULTS Peripherin/rds protein levels in NMP homozygotes were approximately 60% of wild-type levels. Western blot and immunoprecipitation analyses confirmed normal biochemical properties of the NMP protein when compared with wild-type peripherin/rds. Immunohistochemistry demonstrated appropriate localization of transgenic peripherin/rds protein to the disc rim region of photoreceptor outer segments. Total peripherin/rds levels in the retina were modulated by crossing NMP transgenic mice into different rds genetic backgrounds. A positive correlation was observed between peripherin/rds expression levels and the structural and functional integrity of photoreceptor outer segments. Overexpression of peripherin/rds caused no detectable adverse effects on rod or cone structure and function. CONCLUSIONS These findings may have significant implications regarding therapeutic intervention in peripherin/rds-associated retinal diseases.