Analysis of the Ush2a Gene in Medaka Fish (Oryzias latipes)

José L. Mullor,Javier U. Chicote,Ana V. Sánchez-Sánchez,Filippo Del Bene,Elena Aller,Gema García-García,Antonio García-España,Marina Piquer-Gil,Manuel Díaz-Llopis,Patricia Udaondo,Laura Cavallé,José M. Millán

doi:10.1371/journal.pone.0074995

Abstract

Patients suffering from Usher syndrome (USH) exhibit sensorineural hearing loss, retinitis pigmentosa (RP) and, in some cases, vestibular dysfunction. USH is the most common genetic disorder affecting hearing and vision and is included in a group of hereditary pathologies associated with defects in ciliary function known as ciliopathies. This syndrome is clinically classified into three types: USH1, USH2 and USH3. USH2 accounts for well over one-half of all Usher cases and mutations in the USH2A gene are responsible for the majority of USH2 cases, but also for atypical Usher syndrome and recessive non-syndromic RP. Because medaka fish (Oryzias latypes) is an attractive model organism for genetic-based studies in biomedical research, we investigated the expression and function of the USH2A ortholog in this teleost species. Ol-Ush2a encodes a protein of 5.445 aa codons, containing the same motif arrangement as the human USH2A. Ol-Ush2a is expressed during early stages of medaka fish development and persists into adulthood. Temporal Ol-Ush2a expression analysis using whole mount in situ hybridization (WMISH) on embryos at different embryonic stages showed restricted expression to otoliths and retina, suggesting that Ol-Ush2a might play a conserved role in the development and/or maintenance of retinal photoreceptors and cochlear hair cells. Knockdown of Ol-Ush2a in medaka fish caused embryonic developmental defects (small eyes and heads, otolith malformations and shortened bodies with curved tails) resulting in late embryo lethality. These embryonic defects, observed in our study and in other ciliary disorders, are associated with defective cell movement specifically implicated in left-right (LR) axis determination and planar cell polarity (PCP).

Highlights

Usher syndrome (USH) is the most frequent genetic cause of combined deafness and blindness causing hearing loss, retinitis pigmentosa (RP) and, in some cases, vestibular dysfunction
USH2 is characterized by moderate non-progressive hearing loss without vestibular dysfunction and USH3 is distinguished from USH1 and USH2 by the progressive nature of its hearing loss [3]
By PCR amplification of embryonic and ocular cDNA based on these predictions followed by sequencing, the Ol-Ush2a gene was characterized

Summary

Introduction

Usher syndrome (USH) is the most frequent genetic cause of combined deafness and blindness causing hearing loss, retinitis pigmentosa (RP) and, in some cases, vestibular dysfunction. Usher syndrome type I is the most severe form with profound congenital deafness and vestibular dysfunction. USH2 accounts for well over one-half of all Usher cases and up to date, 3 genes are known to be involved in the pathogenesis of this clinical form: USH2A, GPR98 and DFNB31 [4,5,6,7]. Mutations in the USH2A gene are responsible for the majority of USH2 cases [8,9,10] and are responsible for atypical Usher syndrome and recessive non-syndromic RP [11,12]. The short isoform_a, reported to be 5 kb, encoding a protein of 170 kDa; and the long isoform_b, that expands the length of coding sequence to 15 kb, encoding a 600 kDa protein [5]

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