Abstract
Purpose: :
The developmental specialization and functional activity of both retinal photoreceptor cells and mechanosensory hair cells of the inner ear are controlled by many of the same molecules. This common mechanism of regulation is underscored by the pathology of Usher syndrome (USH), a heterogeneous disease characterized by combined vision and hearing loss. We are investigating the roles and interactions of USH genes to understand how they function together to regulate photoreceptor and sensory hair cell development and physiology.
Methods: :
We cloned zebrafish othologues of USH1C, USH2A and USH2C and used cRNA probes to characterize their expression patterns. We injected antisense morpholino oligonucleotides designed to knock down expression in developing embryos and larvae. TUNEL labeling and vital staining of apoptotic cells were used to assay retinal cell death, and antibodies were used to characterize the structural integrity of photoreceptors. We assayed visual function in live larvae with the optokinetic response test.
Results: :
All three genes are expressed in the neural retina and the otic epithelium from late segmentation stages through early larval stages. Transcripts are also present in the adult retina. Knock–down of zebrafish Ush2a by morpholino injection results in swimming defects in young larvae similar to those seen in ush1b (myo7a) mutant fish. We also observe severely reduced visual function, compromised photoreceptor cell integrity and increased TUNEL labeling in the retina within the first week of development.
Conclusions: :
Zebrafish USH orthologues are expressed in tissues consistent with the human disease pathology and at time points that suggest a developmental role in addition to one of maintenance. Depletion of functional Ush2a (Usherin) protein results in loss of visual function, compromised photoreceptor structural integrity and retinal cell death, even in young animals.
Keywords: genetics • retinal degenerations: hereditary • proteins encoded by disease genes