Disruption of CEP290 microtubule/membrane-binding domains causes retinal degeneration
Theodore G. Drivas, … , Erika L.F. Holzbaur, Jean Bennett
Theodore G. Drivas, … , Erika L.F. Holzbaur, Jean Bennett
Published September 24, 2013
Citation Information: J Clin Invest. 2013;123(10):4525-4539. https://doi.org/10.1172/JCI69448.
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Research Article Cell biology

Disruption of CEP290 microtubule/membrane-binding domains causes retinal degeneration

Abstract

Mutations in the gene centrosomal protein 290 kDa (CEP290) cause an array of debilitating and phenotypically distinct human diseases, ranging from the devastating blinding disease Leber congenital amaurosis (LCA) to Senior-Løken syndrome, Joubert syndrome, and the lethal Meckel-Gruber syndrome. Despite its critical role in biology and disease, very little is known about CEP290’s function. Here, we have identified 4 functional domains of the protein. We found that CEP290 directly binds to cellular membranes through an N-terminal domain that includes a highly conserved amphipathic helix motif and to microtubules through a domain located within its myosin-tail homology domain. Furthermore, CEP290 activity was regulated by 2 autoinhibitory domains within its N and C termini, both of which were found to play critical roles in regulating ciliogenesis. Disruption of the microtubule-binding domain in a mouse model of LCA was sufficient to induce significant deficits in cilium formation, which led to retinal degeneration. These data implicate CEP290 as an integral structural and regulatory component of the cilium and provide insight into the pathological mechanisms of LCA and related ciliopathies. Further, these data illustrate that disruption of particular CEP290 functional domains may lead to particular disease phenotypes and suggest innovative strategies for therapeutic intervention.

Authors

Theodore G. Drivas, Erika L.F. Holzbaur, Jean Bennett

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Figure 11

rd16 mouse fibroblasts are deficient in primary cilium formation.

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rd16 mouse fibroblasts are deficient in primary cilium formation. (A)...
(A) Fluorescent microscopy images of serum-starved WT and rd16 primary dermal fibroblasts stained for acetylated α-tubulin (red) to identify primary cilia and stained with DAPI (blue). Scale bars: 5 μm. (B) Average cilium length of serum-starved WT and rd16 primary dermal fibroblasts. Quantification was based on separate experiments on fibroblasts coming from 3 different animals per genotype. At least 50 cilia were measured per experiment, and a total of 400 cilia were measured for both the WT and rd16 fibroblasts. Data are presented as mean ± SD, n = 5. (C) Fluorescence microscopy fields of WT and rd16 primary dermal fibroblasts stained for acetylated α-tubulin (red) to identify primary cilia and stained with DAPI (blue). Fibroblasts were grown in medium with (fed) or without (starved) serum. Arrowheads indicate primary cilia. Scale bars: 10 μm. (D) Percentage of WT and rd16 primary dermal fibroblasts that form primary cilia in serum-fed and serum-starved conditions. Quantification was based on separate experiments on fibroblasts coming from 3 different animals per genotype. At least 100 cells were counted per experiment. Data are presented as mean ± SD, n = 5. *P < 0.05.