Novel truncating variants inCTNNB1cause familial exudative vitreoretinopathy

Abstract

BackgroundFamilial exudative vitreoretinopathy (FEVR) is an inheritable blinding disorder with clinical and genetic heterogeneity. Heterozygous variants in theCTNNB1gene have been reported to cause FEVR. However, the pathogenic basis ofCTNNB1-associated FEVR has not been fully explored.MethodsWhole-exome sequencing was performed on the genomic DNA of probands. Dual-luciferase reporter assay, western blotting and co-immunoprecipitation were used to characterise the impacts of variants. Quantitative real-time PCR, EdU (5-ethynyl-2′-deoxyuridine) incorporation assay and immunocytochemistry were performed on the primary human retinal microvascular endothelial cells (HRECs) to investigate the effect ofCTNNB1depletion on the downstream genes involved in Norrin/β-catenin signalling, cell proliferation and junctional integrity, respectively. Transendothelial electrical resistance assay was applied to measure endothelial permeability. Heterozygous endothelial-specificCtnnb1-knockout mouse mice were generated to verify FEVR-like phenotypes in the retina.ResultsWe identified two novel heterozygous variants (p.Leu103Ter and p.Val199LeufsTer11) and one previously reported heterozygous variant (p.His369ThrfsTer2) in theCTNNB1gene. These variants caused truncation and degradation of β-catenin that reduced Norrin/β-catenin signalling activity. Additionally, knockdown (KD) ofCTNNB1in HRECs led to diminished mRNA levels of Norrin/β-catenin targeted genes, reduced cell proliferation and compromised junctional integrity. The Cre-mediated heterozygous deletion ofCtnnb1in mouse endothelial cells (ECs) resulted in FEVR-like phenotypes. Moreover, LiCl treatment partially rescued the defects inCTNNB1-KD HRECs and EC-specificCtnnb1heterozygous knockout mice.ConclusionOur findings reinforced the current pathogenesis of Norrin/β-catenin for FEVR and expanded the causative variant spectrum ofCTNNB1for the prenatal diagnosis and genetic counselling of FEVR.