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Usher syndrome (USH) is the most common cause of combined deaf-blindness in man. The hearing loss can be partly compensated by providing patients with hearing aids or cochlear implants, but the loss of vision is currently untreatable. In general, mutations in the USH2A gene are the most frequent cause of USH explaining up to 50% of all patients worldwide. The first deep-intronic mutation in the USH2A gene (c.7595-2144A>G) was reported in 2012, leading to the insertion of a pseudoexon (PE40) into the mature USH2A transcript. When translated, this PE40-containing transcript is predicted to result in a truncated non-functional USH2A protein. In this study, we explored the potential of antisense oligonucleotides (AONs) to prevent aberrant splicing of USH2A pre-mRNA as a consequence of the c.7595-2144A>G mutation. Engineered 2'-O-methylphosphorothioate AONs targeting the PE40 splice acceptor site and/or exonic splice enhancer regions displayed significant splice correction potential in both patient derived fibroblasts and a minigene splice assay for USH2A c.7595-2144A>G, whereas a non-binding sense oligonucleotide had no effect on splicing. Altogether, AON-based splice correction could be a promising approach for the development of a future treatment for USH2A-associated retinitis pigmentosa caused by the deep-intronic c.7595-2144A>G mutation.

Ciliopathies are Mendelian disorders caused by dysfunction of cilia, ubiquitous organelles involved in fluid propulsion (motile cilia) or signal transduction (primary cilia). Retinal dystrophy is a common phenotypic characteristic of ciliopathies since photoreceptor outer segments are specialized primary cilia. These ciliary structures heavily rely on intracellular minus-end directed transport of cargo, mediated at least in part by the cytoplasmic dynein 1 motor complex, for their formation, maintenance and function. Ninein-like protein (NINL) is known to associate with this motor complex and is an important interaction partner of the ciliopathy-associated proteins lebercilin, USH2A and CC2D2A. Here, we scrutinize the function of NINL with combined proteomic and zebrafish in vivo approaches. We identify Double Zinc Ribbon and Ankyrin Repeat domains 1 (DZANK1) as a novel interaction partner of NINL and show that loss of Ninl, Dzank1 or both synergistically leads to dysmorphic photoreceptor outer segments, accumulation of trans-Golgi-derived vesicles and mislocalization of Rhodopsin and Ush2a in zebrafish. In addition, retrograde melanosome transport is severely impaired in zebrafish lacking Ninl or Dzank1. We further demonstrate that NINL and DZANK1 are essential for intracellular dynein-based transport by associating with complementary subunits of the cytoplasmic dynein 1 motor complex, thus shedding light on the structure and stoichiometry of this important motor complex. Altogether, our results support a model in which the NINL-DZANK1 protein module is involved in the proper assembly and folding of the cytoplasmic dynein 1 motor complex in photoreceptor cells, a process essential for outer segment formation and function.

Over the last decade, huge progress has been made in the understanding of the molecular mechanisms underlying inherited retinal dystrophy (IRD), as well as in the development and implementation of novel therapies, especially in the field of gene therapy. The use of mutant animal models, either naturally occurring or generated by genetic modification, have contributed greatly to our knowledge on IRD. Yet, these mutant animal models do not always mimic the retinal phenotype that is observed in humans with mutations in the orthologous gene, often due to species-specific characteristics of the retina, and/or diverse functions of the gene products in different species. In this manuscript, we compare general and ocular characteristics of a series of widely used vertebrate animal models, i.e. zebrafish, chicken, rodents, cats, dogs, sheep, pigs and monkeys, in terms of genetic architecture and sequence homology, methods to modify genomes, anatomy of the eye, and structural details of the retina. Furthermore, we present an overview of mutant vertebrate animal models that have been used to study or develop treatments for the various genetic subtypes of IRD, and correlate the suitability of these models to the specific characteristics of each animal. Herewith, we provide tools that will help to select the most suitable animal model for specific research questions on IRDs in the future, and thereby assist in an optimal use of animals and resources to further increase our understanding of inherited retinal dystrophies, and develop novel treatments for these disorders., (Copyright © 2015 Elsevier Ltd. All rights reserved.)