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Erik de Vrieze, Ralph Slijkerman, Margo Dona, Sanne Broekman, Lisette Hetterschijt, Theo Peters, Hannie Kremer, Erwin van Wijk; Contribution of autophagy to Usher syndrome pathogenesis. Invest. Ophthalmol. Vis. Sci. 2017;58(8):2486.
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© ARVO (1962-2015); The Authors (2016-present)
Usher syndrome (USH) is the most common cause of hereditary deaf-blindness. USH patients are born with congenital hearing impairment, and suffer from progressive vision loss (retinitis pigmentosa, RP), a combination that puts them at risk for social isolation and loss of independence. Mutations in the USH2A gene are the most frequent cause of USH, explaining up to 50% of all cases. Mutations in USH2A can also result in non-syndromic RP. Currently, virtually nothing is known about the pathogenesis of USH2A-associated RP.
We used complementary proteomics techniques to identify novel interaction partners of USH2A. Using CRISPR/Cas9-mediated genome editing, we generated an ush2a zebrafish knockout model, carrying a protein-truncating mutation in exon 13. Guided by our proteomic studies, we deep-phenotyped our ush2a-/- zebrafish.
We identified and validated interactions between multiple subunits of the Cop9 signalosome (CSN) and members of the USH protein complex. CSN subunit 8 (COPS8) is a direct interactor of USH2A. CSN/COPS8 is well documented to play a role in two proteostatic pathways: ubiquitin proteasome system (UPS) and autophagy. Immunohistochemistry showed that Cops8 localization in the photoreceptor overlaps with Ush2a, allowing interactions in vivo. Phenotypic analyses of the ush2a-/- retina showed increased levels of photoreceptor apoptosis, as well as an increase in autophagosomes. However, we did not find evidence of increased activity of the UPS system. We also identified mislocalization of rhodopsin-containing transport vesicles. Surprisingly, ER stress (protein-accumulation in the endoplasmic reticulum) appears diminished.
Using zebrafish as a model organism, we have identified that elevated levels of autophagy might be the pathogenic mechanism underlying USH2A-associated retinal degeneration. Whether this is a direct consequence of misregulated autophagy (through CSN), or is a response to the mislocalized transport vesicles, remains to be established. Both prolonged elevation of autophagy and activation of mislocalized photopigments can lead to the observed apoptosis of photoreceptors. The pathways that we have identified explain the slow progressive nature of the retinal degeneration in USH2A patients. This brings us closer to understanding the pathogenesis of USH2A-associated RP, which is also important for the development of future therapies.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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