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JC Corral Serrano, Stef Letteboer, Alex Garanto, Ronald Roepman, Rob W.J. Collin; Investigating the role of C2orf71 in the pathogenesis of retinitis pigmentosa. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4643.
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© ARVO (1962-2015); The Authors (2016-present)
Mutations in C2orf71 are causative for autosomal-recessive retinitis pigmentosa. Although the C2orf71 protein is thought to exert its function in the connecting cilium of photoreceptor cells, its exact role in the retina remains elusive. To gain insight into the physiological function of C2orf71, we conducted a series of experiments involving patient-derived fibroblast cells, immunohistochemical analysis in mouse retinas, and protein-protein interaction studies.
Fibroblasts from a patient with a homozygous C2orf71 mutation, and those of age-matched controls were grown upon serum starvation conditions to induce cilia development. For each cell line, the total number of ciliated cells were counted and the lengths of the cilia were measured. For the localization studies, mouse retina sections were stained with antibodies against C2orf71 and ciliary markers. To find binary interacting partners of C2orf71, we performed a yeast two-hybrid screen of two retinal cDNA libraries as well as a dedicated array of 150 known ciliary proteins using C2orf71 as a bait.
Fibroblasts from a patient homozygously carrying a mutation in C2orf71 did not show a significant difference in the number of ciliated cells nor the average cilium length when compared to control fibroblasts. In the mouse retina, C2orf71 localizes to the connecting cilium of photoreceptor cells, as evidenced by a clear overlapping staining with the ciliary marker acetylated tubulin. A preliminary assessment of the yeast-two-hybrid data revealed several potentially interacting proteins, that are mainly known to be involved in ciliogenesis or ciliary maintenance.
Our results show that mutations in C2orf71 do not affect the structure of the cilium in fibroblast cells, suggesting that the function of C2orf71 may be restricted to photoreceptor cells. The localization of C2orf71 to the connecting cilium in mouse photoreceptors, and its potential affinity to multiple proteins involved in cilium biogenesis and/or homeostasis indicate that C2orf71 exerts its function specifically in the photoreceptor sensory cilium. Our current validation and functional evaluation of the C2orf71 interactome may unveil the details of how gene mutations compromise ciliary function, and thereby result in autosomal recessive retinitis pigmentosa.
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