Abstract
Purpose :
Primary cilia are thin microtubule-based projections that extend from the plasma membrane of quiescent cells. Photoreceptor cells contain a specialized primary cilium called the outer segment, which is responsible for light capture. In general, human mutations present in core ciliary components cause syndromic diseases collectively termed ciliopathies. Interestingly, a subset of these mutations leads to non-syndromic retinal dystrophy. In this study, we identified a human patient with late-onset retinitis pigmentosa caused by a novel frameshift mutation resulting in early truncation of the CEP162 protein. CEP162 is localized to the distal-end of centrioles and required for ciliogenesis. To understand its underlying role in retinal dystrophy, we investigate how the human CEP162 mutation affects protein localization and cilia formation in patient derived human fibroblasts and mouse photoreceptors.
Methods :
Control and patient derived human fibroblasts were analyzed for CEP162 mRNA expression, non-sense mediated decay, CEP162 protein stability and localization, and primary cilia formation and structure. Additionally, CEP162 staining was performed on mouse retinal cross-sections.
Results :
Our analysis of the patient's fibroblast revealed that while truncated mutant CEP162 protein was expressed, it was not localized to the centrosomes. Absence of CEP162 from the basal bodies resulted in reduced ciliation in the patient fibroblasts compared to control. Interestingly, in patient fibroblasts CP110, a negative regulator of ciliogenesis, accumulated at the basal body. Further analysis revealed that although CEP290 was recruited to the basal body polyglutamylated tubulin was absent suggesting that ciliogenesis was halted at the stage of axoneme extension prior to transition zone assembly. Correlative light and scanning electron microscopy (CL-SEM) also showed stalled ciliary structures in the patient fibroblast compared to the control. Immunohistochemistry from mouse retinal cross-sections revealed endogenous CEP162 is localized to the distal end of the basal bodies at the base of the outer segment.
Conclusions :
Our results indicate a novel role for CEP162 mediated axoneme extension, prior to transition zone assembly, as well as establish CEP162 as a retinal dystrophy gene.
This is a 2021 ARVO Annual Meeting abstract.