Abstract:

OBJECTIVES: To identify gene mutations in patients with dystroglycanopathy and prove pathogenicity of those mutations using an in vitro cell assay.
METHODS: We performed whole-exome sequencing on 20 patients, who were previously diagnosed with dystroglycanopathy by immunohistochemistry and/or Western blot analysis. We also evaluated pathogenicity of identified mutations for phenotypic recovery in a DAG1-knockout haploid human cell line transfected with mutated DAG1 complementary DNA.
RESULTS: Using exome sequencing, we identified compound heterozygous missense mutations in DAG1 in a patient with asymptomatic hyperCKemia and pathologically mild muscular dystrophy. Both mutations were in the N-terminal region of α-dystroglycan and affected its glycosylation. Mutated DAG1 complementary DNAs failed to rescue the phenotype in DAG1-knockout cells, suggesting that these are pathogenic mutations.
CONCLUSION: Novel mutations in DAG1 are associated with asymptomatic hyperCKemia with hypoglycosylation of α-dystroglycan. The combination of exome sequencing and a phenotype-rescue experiment on a gene-knockout haploid cell line represents a powerful tool for evaluation of these pathogenic mutations.

日本語要旨:

ジストログリカノパチーが疑われる２０例に対して全エクソーム解析を行い、高CK血 症１例にDAG1遺伝子の複合ヘテロ接合変異を見いだした。DAG1欠損細胞に変異遺伝子を導入したところ、細胞の表現型の回復が見られなかったため、これらの変異が病因性を持つことが証明された。この遺伝子欠損細胞を用いる遺伝子変異証明実験は、様々な遺伝子変異に広く利用できる可能性が考えられた。