Molecular consequences of BEST1 gene mutations in canine multifocal retinopathy predict functional implications for human bestrophinopathies.
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ABSTRACT: Purpose
Bestrophin-1 gene (BEST1) mutations are responsible for a broad spectrum of human retinal phenotypes, jointly called bestrophinopathies. Canine multifocal retinopathy (cmr), caused by mutations in the dog gene ortholog, shares numerous phenotypic features with human BEST1-associated disorders. The purpose of this study was the assessment of molecular consequences and pathogenic outcomes of the cmr1 (C(73)T/R(25)X) premature termination and the cmr2 (G(482)A/G(161)D) missense mutation of the canine model compared with the C(87)G/Y(29)X mutation observed in human patients.Methods
Dogs carrying the BEST1 mutation were introduced into a breeding colony and used to produce either carrier or affected offspring. Eyes were collected immediately after euthanatization at the disease-relevant ages and were harvested for expression studies. In parallel, an in vitro cell culture model system was developed and compared with in vivoResults
Results
The results demonstrate that cmr1 and human C(87)G-mutated transcripts bypass the nonsense-mediated mRNA decay machinery, suggesting the AUG proximity effect as an underlying transcriptional mechanism. The truncated protein, however, is not detectable in either species. The in vitro model accurately recapitulates transcriptional and translational expression events observed in vivo and, thus, implies loss of bestrophin-1 function in cmr1-dogs and Y(29)X-affected patients. Immunofluorescence microscopy of cmr2 mutant showed mislocalization of the protein.Conclusions
Molecular evaluation of cmr mutations in vivo and in vitro constitutes the next step toward elucidating genotype-phenotype interactions concerning human bestrophinopathies and emphasizes the importance of the canine models for studying the complexity of the BEST1 disease mechanism.
SUBMITTER: Guziewicz KE
PROVIDER: S-EPMC3175949 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:To report the ocular phenotype in patients with autosomal recessive bestrophinopathy and carriers, and to describe novel BEST1 mutations.Patients with clinically suspected and subsequently genetically proven autosomal recessive bestrophinopathy underwent full ophthalmic examination and investigation with fundus autofluorescence imaging, spectral domain optical coherence tomography, electroretinography, and electrooculography. Mutation analysis of the BEST1 gene was performed through direct Sanger sequencing.Five affected patients from four families were identified. Mean age was 16 years (range, 6-42 years). All affected patients presented with reduced visual acuity and bilateral, hyperautofluorescent subretinal yellowish deposits within the posterior pole. Spectral domain optical coherence tomography demonstrated submacular fluid and subretinal vitelliform material in all patients. A cystoid maculopathy was seen in all but one patient. In 1 patient, the location of the vitelliform material was seen to change over a follow-up period of 3 years despite relatively stable vision. Visual acuity and fundus changes were unresponsive to topical and systemic carbonic anhydrase inhibitors and systemic steroids. Carriers had normal ocular examinations including normal fundus autofluorescence. Three novel mutations were detected.Three novel BEST1 mutations are described, suggesting that many deleterious variants in BEST1 resulting in haploinsufficiency are still unknown. Mutations causing autosomal recessive bestrophinopathy are mostly located outside of the exons that usually harbor vitelliform macular dystrophy-associated dominant mutations.
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Project description:PURPOSE: To analyze functional and clinical data of Best vitelliform macular dystrophy (VMD) patients with mutations in the BEST1 gene. METHODS: Best VMD patients with BEST1 mutations were evaluated prospectively regarding age, age of onset, best-corrected visual acuity (BCVA), fundus autofluorescence, fluorescein angiography, optical coherence tomography, and electro-oculography. Mutations in BEST1 were established by direct sequencing. RESULTS: Forty-six eyes of 23 patients (10 male, 13 female) were included in the study. We identified nine different BEST1 mutations (3/9 novel), in ten unrelated families. The age of patients ranged between 3 and 75 years; age of onset varied between 2 and 67 years. BCVA ranged between 20/20 and 20/200. On the basis of fundus biomicroscopy with direct illumination, using one widely accepted classification, the macular lesions could be counted as follows: 1. no lesion (normal fovea): eight eyes, five patients carrying a mutation on the BEST1 gene; 2. previtelliform lesions: six eyes, three affected patients; 3. vitelliform lesions: four eyes, two affected patients; 4. pseudohypopyon: three eyes, three affected patients; 5. vitelliruptive lesions (scrambled egg aspect with dispersion of the vitelliform material without sign of atrophy or fibrosis): ten eyes, six affected patients; 6. atrophic lesions (atrophy with or without residual dispersed material): seven eyes, five patients; 7. fibrotic lesions: eight eyes, five patients. Two patients presented unilateral Best VMD. Both eyes of two patients presented multifocal Best VMD features on fundus examination. Six eyes of four patients have been treated for choroidal neovascularization by thermic photocoagulation [one eye], photodynamic therapy [three eyes], and intravitreal ranibizumab injection [two eyes]. Comparison of interfamilial and intrafamilial clinical data between patients did not reveal differences in age, BCVA, and stage of the disease as evaluated by fundus autofluorescence, fluorescein angiography, and optical coherence tomography (p>0.05). Mean BCVA impairment showed a statistically significant correlation to a more advanced stage of the disease (p<0.001). CONCLUSIONS: BEST1 mutations were not correlated with the severity of the functional and clinical data in the Best VMD patients examined.
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