Leber congenital amaurosis (LCA) causes blindness or severe visual impairment at or within a few months of birth. Here we show, using homozygosity mapping, that the LCA5 gene on chromosome 6q14, which encodes the previously unknown ciliary protein lebercilin, is associated with this disease. We detected homozygous nonsense and frameshift mutations in LCA5 in five families affected with LCA. In a sixth family, the LCA5 transcript was completely absent. LCA5 is expressed widely throughout development, although the phenotype in affected individuals is limited to the eye. Lebercilin localizes to the connecting cilia of photoreceptors and to the microtubules, centrioles and primary cilia of cultured mammalian cells. Using tandem affinity purification, we identified 24 proteins that link lebercilin to centrosomal and ciliary functions. Members of this interactome represent candidate genes for LCA and other ciliopathies. Our findings emphasize the emerging role of disrupted ciliary processes in the molecular pathogenesis of LCA.

The Crumbs proteins (CRBs) are transmembrane proteins, homologous to Drosophila Crumbs, with a key role in defining the apical membrane domain in photoreceptors as well as in embryonic epithelia. Crumbs proteins are conserved between species and their intracellular domains are involved in organizing a conserved macromolecular protein scaffold with important roles in cell polarity as well as morphogenesis and maintenance of the retina. Mutations in the gene encoding human CRB1, the first one identified out of the three human orthologs, have been associated with a number of retinal dystrophies including Leber amaurosis and retinitis pigmentosa type 12. Although no other mammalian Crumbs complex members as of yet have been associated with retinal degeneration, disruption of different zebrafish and fruitfly orthologs can lead to various retinal defects. The core Crumbs complex localizes apical to the outer limiting membrane, where photoreceptors and Müller glia contact each other. Correct functioning of Crumbs ensures adhesion between these cells by an unknown mechanism. This review summarizes the current view on the composition and function of the Crumbs prsotein complex in the mammalian retina. Recently, a number of new members of the Crumbs protein complex have been identified. These include most members of the membrane palmitoylated protein family (MPP), involved in assembly of macromolecular protein complexes. Some components of the complex are found to exert a function in the photoreceptor synapses and/or at the region of the connecting cilium. Studies using polarized cell cultures or model organisms, like Drosophila and zebrafish, suggest important links of the Crumbs protein complex to several biological processes in the mammalian eye, including retinal patterning, ciliogenesis and vesicular transport.

These data imply that MPP4 and -5 have a role in photoreceptor polarity and, by association with CRB1, pinpoint the cognate genes as functional candidate genes for inherited retinopathies.

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The highly ordered distribution of neurons is an essential feature of a functional mammalian retina. Disruptions in the apico-basal polarity complexes at the outer limiting membrane (OLM) of the retina are associated with retinal patterning defects in vertebrates. We have analyzed the binding repertoire of MPP5/Pals1, a key member of the apico-basal Crumbs polarity complex, that has functionally conserved counterparts in zebrafish (nagie oko) and Drosophila (Stardust). We show that MPP5 interacts with its MAGUK family member MPP1/p55 at the OLM. Mechanistically, this interaction involves heterodimerization of both MAGUK modules in a directional fashion. MPP1 expression in the retina throughout development resembles the expression of whirlin, a multi-PDZ scaffold protein and an important organizer in the Usher protein network. We demonstrate that both proteins interact strongly by both a classical PDZ domain-to-PDZ binding motif (PBM) mechanism, and a mechanism involving internal epitopes. MPP1 and whirlin colocalize in the retina at the OLM, at the outer synaptic layer and at the basal bodies and the ciliary axoneme. In view of the known roles of the Crumbs and Usher protein networks, our findings suggest a novel link of the core developmental processes of actin polymerization and establishment/maintenance of apico-basal cell polarity through MPP1. These processes, essential in neural development and patterning of the retina, may be disrupted in eye disorders that are associated with defects in these protein networks.