Functional characterization of variants found in Japanese patients with hereditary hemorrhagic telangiectasia

<jats:title>Abstract</jats:title><jats:p>Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant form of vascular dysplasia. Genetic diagnosis is made by identifying loss‐of‐function variants in genes, such as <jats:italic>ENG</jats:italic> and <jats:italic>ACVRL1</jats:italic>. However, the causal mechanisms of various variants of unknown significance remains unclear. In this study, we analyzed 12 Japanese patients from 11 families who were clinically diagnosed with HHT. Sequencing analysis identified 11 distinct variants in <jats:italic>ACVRL1</jats:italic> and <jats:italic>ENG</jats:italic>. Three of the 11 were truncating variants, leading to a definitive diagnosis, whereas the remaining eight were splice‐site and missense variants that required functional analyses. In silico splicing analyses demonstrated that three variants, c.526‐3C > G and c.598C > G in <jats:italic>ACVRL1</jats:italic>, and c.690‐1G > A in <jats:italic>ENG</jats:italic>, caused aberrant splicing, as confirmed by a minigene assay. The five remaining missense variants were p.Arg67Gln, p.Ile256Asn, p.Leu285Pro, and p.Pro424Leu in <jats:italic>ACVRL</jats:italic> and p.Pro165His in <jats:italic>ENG</jats:italic>. Nanoluciferase‐based bioluminescence analyses demonstrated that these <jats:italic>ACVRL1</jats:italic> variants impaired cell membrane trafficking, resulting in the loss of bone morphogenetic protein 9 (BMP9) signal transduction. In contrast, the <jats:italic>ENG</jats:italic> mutation impaired BMP9 signaling despite normal cell membrane expression. The updated functional analysis methods performed in this study will facilitate effective genetic testing and appropriate medical care for patients with HHT.</jats:p>