Molecular consequences of BEST1 gene mutations in canine multifocal retinopathy predict functional implications for human bestrophinopathies

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.

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 vivo

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.

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.