CLARIN 1 RETINAL FUNCTION AND THERAPEUTIC IMPLICATIONS FOR USH3

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Project Summary/Abstract Mutations in the Clarin1 (CLRN1) gene cause Usher syndrome type 3 (USH3), a devastating orphan disease leading to combined blindness and deafness in humans. Despite its very low levels of expression, the lack of CLRN1 results in progressive degeneration of rod and cone photoreceptors and cochlear hair cells. There is no treatment currently available to prevent vision loss. The lack of animal models that display a retinal phenotype has been a major barrier to understanding the USH3 retinal pathophysiology and developing therapies to prevent the progressive degeneration of the photoreceptor cells. In recent studies, we and others reported the surprising discovery that CLRN1 mRNA is enriched in retinal Müller glia across several vertebrate species. Our combined published results challenge a long-standing photoreceptor-driven degenerative mechanism in USH3 and suggest that pathology arises from disruption of key Müller glia-photoreceptor interactions. The significance of CLRN1 expression in Müller glia and the sequence of pathological events culminating in widespread photoreceptor cell loss are unknown. To address these gaps in our knowledge, and test the hypothesis that USH3 is a Müller glia-driven disease affecting photoreceptors, this project proposes a cross-phylogenetic integration approach with newly developed animal models of USH3, zebrafish and pig. In our preliminary data, we show that our zebrafish model lacking clarin1 displays an age-dependent loss of photoreceptors. We will selectively restore or delete CLRN1 expression specifically in Müller glia and determine whether CLRN1 presence/absence alters photoreceptor function, structure, and survival (Aim1). To identify the molecular and cellular basis of USH3 retinal disease in a large animal model, we generated USH3 pigs with heterozygous, biallelic CLRN1 mutations containing distinct insertions/deletions (indels) in exon 1. Many biallelic heterozygous combinations of USH3 mutant pigs have profound hearing loss. A smaller subset shows alterations in the outer retinal laminae detectable by optical coherence tomography and changes in the scotopic electroretinogram. Therefore, we will compare the development of retinal disease across four independent lines of homozygous biallelic USH3 mutant pigs (same indel) to identify the phenotype-driving mutation and specific cellular and molecular changes in Müller glia and photoreceptors (Aim 2). Successful completion of this project will significantly advance the field by providing new animal models and fundamental knowledge on USH3 disease, to ultimately accelerate the development and validation of treatments to prevent blindness.