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D. S. Kasprick, L. M. Ritter, N. Khattree, N. S. Peachey, R. N. Farris, L. Dang, M. Yu, A. R. Bottrell, A. F. X. Goldberg; An Intramembrane Glutamic Acid Governs Peripherin/rds Function for Photoreceptor Disk Morphogenesis. Invest. Ophthalmol. Vis. Sci. 2007;48(13):591.
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© ARVO (1962-2015); The Authors (2016-present)
Genetic defects in peripherin/rds (P/rds) are responsible for a broad diversity of progressive retinal degenerations; however this protein’s normal function at the molecular level, and its role in disease remain uncertain. P/rds is restricted to the rim regions of photoreceptor outer segment (OS) disk membranes, where it interacts with the homologous protein rom-1, to support OS morphogenesis and architecture. A functional asymmetry exists between these homologs in vivo, but the molecular determinants governing their relative efficacies are not known. Here, we examine the significance of an intramembrane glutamic acid residue, conserved in all P/rds proteins, but absent in rom-1 orthologs.
Site-directed mutagenesis was used to substitute a glutamine (for glutamate) into the fourth transmembrane domain of bovine P/rds, and E276Q P/rds was expressed in cultured COS-1 cells and transgenic WT and rds mouse photoreceptors. Western blot, immunoprecipitation, and sedimentation analyses were used to assess protein structure and interactions. Light, fluorescence, and electron microscopy, and electroretinography were used to characterize retinal structure, transgenic protein localization, and photoreceptor ultrastructure, viability, and function.
E276Q P/rds is expressed, assembled, and properly localized in photoreceptor OSs of transgenic mice. In contrast to WT however, this mutant does not rescue the OS structural defects observed in rds mice. Moreover, its expression does not prevent the retinal degeneration that occurs as a consequence of OS disruption.
These results demonstrate that E276 plays an essential role for P/rds support of disk morphogenesis, and provide a molecular rationale for asymmetry in P/rds and rom-1 function.
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