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J. Zuo, J. Liu, T. Yamashita, R. Griffith, A. Romeo, S.J. Bowne, S.P. Daiger, L.S. Sullivan, M.E. C. Fitzgerald; RP1L1 Is Required for Appropriate Outer Segment Formation . Invest. Ophthalmol. Vis. Sci. 2006;47(13):4749.
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© ARVO (1962-2015); The Authors (2016-present)
Retinitis pigmentosa 1 (RP1) is a microtubule–associated protein specific for photoreceptors. An RP1–like protein, RP1L1, shares homology with RP1 only at its N–terminus to the DCX domain known as a microtubule–binding domain, and an adjacent 34–amino acid segment. Both RP1 and RP1L1 are specifically expressed in photoreceptors in similar temporal patterns. We hypothesized that RP1L1 is a microtubule–associated protein and involved in outer segment formation. To investigate the function of RP1L1, and potential interactions between RP1 and RP1L1, we created and characterized Rp1L1 knockout mice.
We raised polyclonal antibodies against the C–terminal recombinant Rp1L1 protein in rabbits. Using ES cells, we targeted Rp1L1 in mice by deleting exons 2, 3, and a large portion of exon 4. Eyes from Rp1L1 knockout mice and wildtype littermates were collected at 1, 3, and 6 months of age for TEM, Western blot, and immunostaining analyses.
Western blot analysis using Rp1L1 antibodies demonstrated that Rp1L1 encodes a 200–kD protein in mouse retinas. Genomic Southern blot and real–time RT–PCR analyses confirmed the deletion of correct Rp1L1 exons and the absence of retinal Rp1L1 mRNA in homozygous mutant mice. Rp1L1–/– mice were viable. Light microscopy and TEM analyses of retinas of F2 mice at 1 and 6 months revealed that a small percentage of photoreceptors of Rp1L1–/– mice displayed abnormal outer segments (OS). Distal and middle portions of individual OS became enlarged, disoriented and disintegrated while surrounded by normal OS; some inner segments and nuclei of photoreceptors that were associated with degenerating OS were also degenerating. At 6 months, only one or two rows of photoreceptors have degenerated in Rp1L1–/– mice.
Preliminary analyses showed that similar to RP1, RP1L1 is required for outer segment formation. The retinas of Rp1L1–/–mice displayed less severe and slightly different phenotypes that those of Rp1–/– mice. Additional experiments are in progress to refine the subcellular localization of RP1L1 and its interactions with RP1.
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