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Bhagwat V Alapure, Katelyn N Robillard, Frank Rigo, Jessie J Guidry, Jennifer J Lentz; Antisense oligonucleotides rescue differentially expressed proteins in the retina of Usher mouse model. Invest. Ophthalmol. Vis. Sci. 2020;61(7):2444.
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Harmonin, a scaffolding protein encoded by the USH1C gene, is expressed in hair cells and photoreceptors. In the ear, harmonin plays an important role in hair cell development and function through interactions with other proteins and the stereocilia; however, its role in the retina is not known. Mutations in harmonin cause Usher syndrome (Usher), the leading genetic cause of deaf-blindness. Mice with the USH1C c.216G>A mutation that causes Type 1 Usher in Acadian populations of the USA and Canada are deaf and have balance and visual dysfunction similar to patients. To understand the effect of the 216A mutation on protein expression in the retina, we performed quantitative discovery-based proteomics of retinal extracts from Usher, wild-type controls (WT), and 216A-targeted antisense oligonucleotide (ASO)-treated Usher mice. We hypothesize that Usher retinas have abnormal protein expression compared with WT controls. Furthermore, we predict that ASO treatment, which improves Ush1c expression and restores visual function in Usher mice, rescues the differentially expressed proteins in retina.
Retinas harvested from Usher, ASO-Usher, and WT mice at various ages and times-post ASO treatment were processed for proteomic analysis using liquid chromatography and mass spectrometry. Proteins were analyzed using Proteome Discoverer 2.3; and confirmed with immunohistochemistry (IHC) and western blot analysis. Visual function was measured using electroretinogram (ERG) analysis.
A total of 3500-4000 proteins were identified in Usher, WT and ASO-treated Usher retinas at 1, 3, 6 and 12 months of age. Of these, 153-245 proteins related to pathways such as oxidative stress and protein transport; were significantly differentially expressed in Usher retinas compared with WT mice. ERGs were significantly reduced in Usher mice at all ages compared with WT mice. Among the differentially expressed proteins in Usher retinas, 7 – 40 proteins, including Superoxide Dismutase (SOD) and Arrestin, returned to wild-type levels with ASO treatment. IHC and western blot confirmed protein levels. These data correlated with significantly improved ERGs in ASO-Usher mice compared to untreated Usher mice.
These data demonstrate that ASO treatment rescues retina specific proteins and visual function in USH1C mice, and suggest a new role for harmonin in the healthy and USH1C retina.
This is a 2020 ARVO Annual Meeting abstract.
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