Retina- and rod-specific knockout of centrosomal protein CEP164

Abstract

Purpose : CEP164 (NPHP15) is a centrosomal protein (1333 amino acids in mouse) located in distal appendages of the basal body and involved in ciliogenesis. Recessive mutations have been shown to be causative of nephronophthisis, Senior-Løken and Bardet-Biedl Syndromes. CEP164 forms a complex with INPP5E, ARL13B and PDE6D in order to target INPP5E to the cilia in IMCD3 cells. CEP164 also plays a role in recruiting TTBK2 to remove the CP110 capping protein to initiate ciliogenesis in mammalian cultured cells. Global knockout of CEP164 is embryonic lethal in mouse. Conditional knockout of CEP164 in multiciliated cells results in loss of airway, ependymal and oviduct multicilia, as well as hydrocephalus and male infertility. We hypothesize that CEP164 plays a role in photoreceptor primary ciliogenesis and ciliary maintenance, so that loss of CEP164 results in retinal degeneration.

Methods : Cep164^f/f mice were donated by Ken-Ichi Takemaru (Stony Brook, N.Y.). Cep164^f/f mice were bred with Six3Cre and iCre75 transgenic lines to create retina- and rod-specific CEP164 knockouts. Electroretinography (ERG) was performed to assess the functional integrity of the knockout retinas. Immunohistochemistry was performed to determine mislocalization or mistrafficking of various outer segment proteins.

Results : CEP164^retKO mice lack connecting cilium, axoneme and outer segment formation. By P21, scotopic and photopic A- and B-wave amplitudes are significantly decreased and photoreceptors show progressive degeneration, as measured by outer nuclear layer (ONL) thickness. CEP164^rodKO mice have normal rod and cone development. By P21, rods show significant decrease in A- and B-wave amplitudes on scotopic ERG, degeneration in the ONL and shortening of the outer segments. CEP164-interacting protein ARL13B also shows mislocalization. By P30, scotopic and photopic ERG are significantly depressed and the ONL is decreased to a single layer of nuclei.

Conclusions : CEP164 is necessary for ciliogenesis in photoreceptors. Lack of outer segment formation likely is the cause of the Leber congenital amaurosis phenotype present in patients with Senior-Løken Syndrome. CEP164 also appears to play a role in proper trafficking of ARL13B to the outer segments. Inability for ARL13B to localize correctly and act as a GEF for ARL3, a cargo displacement factor, may provide a molecular reason for the retinitis pigmentosa phenotype in patients with Bardet-Biedl Syndrome.

This is a 2020 ARVO Annual Meeting abstract.