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H. Khanna, E. Oh, A. Estrada, S. He, A. Karoukis, M. Othman, J. R. Heckenlively, N. Katsanis, A. Swaroop; Interaction of RPGR With Selected BBS Proteins Is Perturbed in Retinal Ciliopathies. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3739.
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© ARVO (1962-2015); The Authors (2016-present)
Mutations in Retinitis Pigmentosa GTPase Regulator (RPGR), a ciliary protein, are associated with a majority of XLRP and simplex cases. However, the function of RPGR is poorly understood. Since retinal degeneration is a typical feature in Bardet-Biedl Syndrome (BBS), a syndromic ciliopathy, we hypothesized that RPGR associates with BBS proteins in the retina and that disruption of such interactions underlies the associated retinal degeneration in XLRP as well as BBS patients.
We performed known-bait and known-prey analysis to assess binary interactions of RPGR with BBS proteins. Macromolecular interactions were determined by co-immunoprecipitation and immunoblot analysis using transfected cell lines and mammalian retinal extracts. BBS patient DNA samples were analyzed for RPGR mutations using standard protocols.
We show that RPGR interacts directly with BBS4 and not with BBS2, BBS5, BBS6, BBS8, or CEP290/NPHP6/BBS12. Full-length BBS4 interacts with the amino-terminal RCC1-like domain of RPGR. Co-immunoprecipitation studies using bovine retinal extracts demonstrate that RPGR exists in macromolecular complex(es) with BBS2, BBS4, BBS6, and CEP290/BBS12. The integrity of the RPGR-BBS complex is perturbed in the Rpgr-knock out, Bbs4 ko, and rd16 (retinal degeneration 16; Cep290/Bbs12 mutation) mice. Sequence analysis of the RPGR gene revealed a missense and a predicted truncation mutation in BBS patients, indicating that RPGR mutations may modify the BBS phenotype in patients.
These results suggest that perturbation of the RPGR-BBS complex(es) due to mutations in either RPGR or BBS may result in misregulation of photoreceptor ciliary transport and retinal degeneration. We propose that analysis of modifier alleles especially in rare genetic diseases is important to determine the mechanism of associated genetic and clinical heterogeneity.
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