Comprehensive genetic analysis reveals the mutational landscape of ABCA4-associated retinal dystrophy in a Chinese cohort

Purpose: To depict the variant profiles of the ABCA4 gene in a large Chinese cohort of patients with ABCA4- associated retinal dystrophy (ABCA4-RD).Methods: We recruited 290 unrelated Chinese patients with ABCA4-RD and did ABCA4 mutational screening by a combination of Sanger sequencing, targeted exome sequencing, entire ABCA4 locus sequencing, and whole genome sequencing (WGS). The pathogenicity of variants was assessed using in silico tools or in vitro splicing assays following the American College of Medical Genetics and Genomics guidelines.Results: Two hundred sixty-eight distinct pathogenic variants were identified, and 57 were novel. In 580 alleles, 22 noncoding region variants outside canonical splice sites and 4 structural variations were found in 44 alleles accounting for 7.6% of all alleles. Bioinformatics analysis showed the complex mechanism of aberrant splicing productsnatural splice site disruption, branch point destruction, and cryptic splice site activation. Correspond-ingly, minigene assays validated the various abnormal splicing products, including exon skipping, exon elon-gation, partial exon deletion, and pseudoexon insertion. WGS identified the first inversion variation in ABCA4.Conclusions: This study systematically depicted the variant profiles of ABCA4 and revealed the missing alleles of patients with ABCA4-RD in a large Chinese cohort. Our findings demonstrated the complexity of molecular diagnosis of Mendelian diseases and the efficiency of WGS for detecting structural variants.