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X-Q Ding, M Nour, A Quiambao, MI Naash; Biochemical and Functional Effects of the Arg172Trp Mutation in Peripherin/rds . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1884.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To investigate the effect of expression of peripherin/rds (P/rds) with the Arg172Trp mutation on the maintenance of morphogenesis and integrity of the photoreceptor outer segments. Methods: Two transgenes have been generated carrying P/rds with: (i) Pro341Gln modification at the C-terminus to allow recognition by the monoclonal antibody 3B6 as a control and (ii) Pro341Gln transgene with Arg172Trp mutation that causes Macular Dystrophy in humans. The transgenes are composed of a 1.3kb IRBP promoter, 1.6kb full-length mouse P/rds cDNA and a 0.9kb SV40 poly A signal. 3B6 antibody recognizes human, bovine, and rat but not murine P/rds. Anatomical, functional and biochemical approaches were performed to determine the effect of the mutation on the photoreceptor cells and to explore the underlying mechanisms. Results: Eleven and thirteen lines were generated for Pro341Gln and Arg172Trp transgenes, respectively. High expressing lines of Pro341Gln and Arg172Trp in +/+, rds-/+ and rds-/- backgrounds were characterized. Histological and electrophysiological analysis of the transgenic mouse retinas showed no degeneration in rods up to 1-year of age. However, a significant reduction in cone photoreceptor cell response in spectral ERG was observed in transgenic mice in 12 and 9 months of age in the +/+ and rds-/+ backgrounds, respectively. Consistent with the functional loss, the number of cone cells was reduced by 30%. This indicates a cone cell specific defect in these transgenic retinas. Western blot analysis confirmed expression of the mutant P/rds in rds-/- background. Co-immunoprecipitation assay showed the normal association of Pro341Gln and Arg172Trp with Rom-1. However, in non-reducing gels, the abundance of P/rds homodimers in Arg172Trp retina in the rds-/- background was significantly reduced when compared to wild-type or Pro341Gln. Studies are currently underway to evaluate the defective role of the Arg172Trp mutation on P/rds and its association with other proteins. Conclusion: Expression of P/rds with Arg172Trp mutation in transgenic mice caused late onset dominant negative effect on cones but not rods. The structural and functional defects in cones are most likely due to the abnormality of the complex formation of the Arg172Trp protein. It is likely that the susceptibility of cone cells to the mutation is related to the specific structural characteristics of the cone outer segments.
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