Exome Sequencing Analysis Identifies Compound Heterozygous Mutation in ABCA4 in a Chinese Family with Stargardt Disease

et al. (2014) Exome Sequencing Analysis Identifies Compound Heterozygous Mutation in ABCA4 in a Chinese Family with Stargardt Disease. PLoS ONE 9(3): e91962. doi:10.1371/journal.pone.0091962 Exome Sequencing Analysis Identifies Compound Heterozygous Mutation in ABCA4 in a Chinese Family with Stargardt Disease Yu Zhou 0 Siyu Tao 0 Hui Chen 0 Lulin Huang 0 Xiong Zhu 0 Youping Li 0 Zhili Wang 0 He Lin 0 Fang Hao 0 Zhenglin Yang 0 Liya Wang 0 Xianjun Zhu 0 Michael G. Anderson, University of Iowa, United States of America 0 1 Sichuan Provincial Key Laboratory for Human Disease Gene Study and Institute of Laboratory Medicine, Sichuan Academy of Medical Sciences and Sichuan Provincial People's Hospital , Chengdu, Sichuan , China , 2 School of Medicine, University of Electronic Science and Technology of China , Chengdu, Sichuan , China , 3 Henan Eye Hospital and Henan Eye Institute, People's Hospital of Zhengzhou University , Zhengzhou, Henan , China , 4 Department of Ophthalmology, Sichuan Academy of Medical Sciences and Sichuan Provincial People's Hospital , Chengdu, Sichuan , China , 5 Laboratory Animal Institute, Sichuan Academy of Medical Sciences and Sichuan Provincial People's Hospital , Chengdu, Sichuan , China , 6 Chengdu Institute of Biology, Chinese Academy of Sciences and Chinese Academy of Sciences Sichuan Translational Medicine Research Hospital , Chengdu, Sichuan , China , 7 Key Laboratory for NeuroInformation of Ministry of Education, University of Electronic Science and Technology of China , Chengdu, Sichuan , China , 8 College of Life Sciences and Engineering, Xinan Jiaotong University , Chengdu, Sichuan , China Stargardt disease is the most common cause of juvenile macular dystrophy. Five subjects from a two-generation Chinese family with Stargardt disease are reported in this study. All family members underwent complete ophthalmologic examinations. Patients of the family initiated the disease during childhood, developing progressively impaired central vision and bilateral atrophic macular lesions in the retinal pigmental epithelium (RPE) that resembled a ''beaten-bronze'' appearance. Peripheral venous blood was obtained from all patients and their family members for genetic analysis. Exome sequencing was used to analyze the exome of two patients II1, II2. A total of 50709 variations shared by the two patients were subjected to several filtering steps against existing variation databases. Identified variations were verified in all family members by PCR and Sanger sequencing. Compound heterozygous variants p.Y808X and p.G607R of the ATP-binding cassette, sub-family A (ABC1), member 4 (ABCA4) gene, which encodes the ABCA4 protein, a member of the ATP-binding cassette (ABC) transport superfamily, were identified as causative mutations for Stargardt disease of this family. Our findings provide one novel ABCA4 mutation in Chinese patients with Stargardt disease. - Funding: This research was supported by the National Natural Science Foundation of China [81025006, 81170883 (Z.Y.), 81271007 (X.Z.)], and Department of Science and Technology of Sichuan Province, China (SZ20120209, Z.Y.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. . These authors contributed equally to this work. Stargardt disease (STGD), which is also known as juvenile macular degeneration, was first reported by Karl Stargardt in 1909. It is one of the most common hereditary retinal dystrophies with an estimated prevalence of at least 1:10,000 [1,2,3]. It presents with a progressive and significant loss in central vision in the first or second decade of life. However, fundus examination is frequently normal early in the course of disease, even when patients already complain of vision loss. At this stage, the clinical diagnosis of Stargardt disease may be missed. Later on, typical fundus manifestations arise, including pigment mottling, frank macular atrophy, a bulls eye maculopathy and fundus flecks in the macular and the perimacular region [4]. However, it should be noted that Stargardt disease presents with highly variable phenotypes. Histologically, Stargardt disease is associated with significant loss in photoreceptor cells and a massive deposition of lipofuscin-like material in the retinal pigment epithelium, which has also been observed in aging human eyes [6,7]. STGD is predominantly inherited as an autosomal recessive trait, although an autosomal dominant form has been also described [5]. Both sexes are equally affected. The STGD gene has been mapped to the short arm of chromosome 1 [8] in a narrow genetic interval, subsequently assigned to band p22.1 [9], now known as ATP-binding cassette, sub-family A (ABC1), member 4 (ABCA4) [10,11]. The gene for an autosomal dominant disorder with a similar phenotype has been reported on chromosome 6 [12]. Autosomal dominant families linked to this locus are at least 50 times less common than families that are consistent with autosomal recessive inheritance [13,14]. Recessively inherited Stargardt disease is likely to be monogenic. Rare cases of STGD or Stargardt-like disease phenotypes have been reported with mutations in PRPH2 [15,16], VMD2 [17], ELOVL4 [5,18,19] and PROM1 [20]. These genes, as well as ABCA4, are also associated with clinically distinct phenotypes including retinitis pigmentosa, cone/rod dystrophy and pattern dystrophy. Known candidate genes for Stargardt disease such as ABCA4 contain many exons and there are hundreds of identified mutations. The cost and time requirement for mutation screening of all coding exons by Sanger sequencing would equal or exceed that of high-throughput next generation sequencing (NGS) analysis. In this study, we applied next generation sequencing technology to identify the disease-causing gene in this family as part of a large cohort study for retinal diseases. Next-generation sequencing, in particular whole-exome sequencing (WES) can now be performed rapidly and at minimal cost, allowing analysis of the coding regions (exome) of the human genome in single individuals or small families, including patients in whom a clear genotypephenotype correlation is absent or for clinically and genetically heterogeneous conditions [21,22,23]. In the present study, disease-associated mutations were identified by WES of the two affected siblings followed by validation in the family affected by Stargardt disease. Our results identified two compound heterozygous disease-segregating mutations, c.C2424G, p.Y808X and c.G1819A, p.G607R, in the ABCA4 gene. To exclude the possibility that these mutations were polymorphisms, DNA samples of 1000 unaffected individuals were also screened for these mutations. Materials and Methods Subjects and Clinical Assessment Study approval was obtained from the Institutional Review Boards of Sichuan Academy of Medical Scienc (...truncated)