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O.L. Moritz, B.M. Tam; Rhodopsin K296E Expressed in Transgenic X. laevis Photoreceptors Resembles a Class II Rhodopsin Mutant . Invest. Ophthalmol. Vis. Sci. 2005;46(13):5191.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: The rhodopsin mutation K296E causes an aggressive form of retinititis pigmentosa (RP). When expressed in cultured cells, the mutant protein constitutively activates transducin. However, in rod photoreceptors, a constitutively active rhodopsin would probably bind arrestin, leading to inactivation. To better understand the effects of K296E rhodopsin on the retina, we expressed this mutant in X. laevis photoreceptors. Methods: Using site–directed mutangenesis, we introduced the K296E mutation into an epitope–tagged form of X. laevis rhodopsin. Mutagenized and non–mutagenized epitope–tagged rhodopsins were expressed in transgenic X. laevis rods. We compared the expression level, localization patterns, and effects on the retina of these gene products by dot blot assays and confocal microscopy in 14–day old animals. Results: K296E epitope tagged rhodopsin was expressed at low levels, and caused aggressive retinal degeneration in many animals. In contrast, control epitope–tagged rhodopsin was expressed at high levels, and did not cause significant retinal degeneration. While the majority of control epitope–tagged rhodopsin was transported to the outer segments, K296E rhodopsin was largely retained in inner segments. A small quantity of K296E rhodopsin was transported to the OS. Conclusions: K296E rhodopsin causes aggressive retinal degeneration in X. laevis. The localization of the mutant protein and low expression level suggests that the gene product is recognized as misfolded by quality control mechanisms of the IS. These results are similar to those we obtained previously using the class II mutant P23H, suggesting that these mutations may cause RP by related mechanisms.
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