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The intronic ABCA 4 c.5461‐10T>C variant, frequently seen in patients with Stargardt disease, causes splice defects and reduced ABCA 4 protein level
Author(s) -
Aukrust Ingvild,
Jansson Ragnhild W.,
Bredrup Cecilie,
Rusaas Hilde E.,
Berland Siren,
Jørgensen Agnete,
Haug Marte G.,
Rødahl Eyvind,
Houge Gunnar,
Knappskog Per M.
Publication year - 2017
Publication title -
acta ophthalmologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.534
H-Index - 87
eISSN - 1755-3768
pISSN - 1755-375X
DOI - 10.1111/aos.13273
Subject(s) - abca4 , stargardt disease , rna splicing , microbiology and biotechnology , exon , biology , fibroblast , splice , polymerase chain reaction , messenger rna , genetics , chemistry , rna , gene , cell culture , phenotype
Abstract Purpose Despite being the third most common ABCA 4 variant observed in patients with Stargardt disease, the functional effect of the intronic ABCA 4 variant c.5461‐10T>C is unknown. The purpose of this study was to investigate the molecular effect of this variant. Methods Fibroblast samples from patients carrying the ABCA 4 variant c.5461‐10T>C were analysed by isolating total RNA , followed by real‐time polymerase chain reaction (RT‐PCR) using specific primers spanning the variant. For detection of ABCA 4 protein, fibroblast samples were lysed and analysed by SDS ‐ PAGE followed by immunoblotting using a monoclonal ABCA 4 antibody. Results The ABCA 4 variant c.5461‐10T>C causes a splicing defect resulting in the reduction of full‐length mRNA in fibroblasts from patients and the presence of alternatively spliced mRNA s where exon 39–40 is skipped. A reduced level of full‐length ABCA 4 protein is observed compared to controls not carrying the variant. Conclusions This study describes the functional effect and the molecular mechanism of the pathogenic ABCA 4 variant c.5461‐10T>C. The variant is functionally important as it leads to splicing defects and a reduced level of ABCA 4 protein.