A comprehensive survey of sequence variation in the ABCA4 (ABCR) gene in Stargardt disease and age-related macular degeneration

Abstract

Stargardt disease (STGD) is a common autosomal recessive maculopathy of early and young-adult onset and is caused by alterations in the gene encoding the photoreceptor-specific ATP-binding cassette (ABC) transporter (ABCA4). We have studied 144 patients with STGD and 220 unaffected individuals ascertained from the German population, to complete a comprehensive, population-specific survey of the sequence variation in the ABCA4 gene. In addition, we have assessed the proposed role for ABCA4 in age-related macular degeneration (AMD), a common cause of late-onset blindness, by studying 200 affected individuals with late-stage disease. Using a screening strategy based primarily on denaturing gradient gel electrophoresis, we have identified in the three study groups a total of 127 unique alterations, of which 90 have not been previously reported, and have classified 72 as probable pathogenic mutations. Of the 288 STGD chromosomes studied, mutations were identified in 166, resulting in a detection rate of approximately 58%. Eight different alleles account for 61% of the identified disease alleles, and at least one of these, the L541P-A1038V complex allele, appears to be a founder mutation in the German population. When the group with AMD and the control group were analyzed with the same methodology, 18 patients with AMD and 12 controls were found to harbor possible disease-associated alterations. This represents no significant difference between the two groups; however, for detection of modest effects of rare alleles in complex diseases, the analysis of larger cohorts of patients may be required.

Figure 1

Evaluation of consequences of intervening sequence alterations: results of RT-PCR analyses (a) and schematic representations (b–f), indicating abnormal splicing at mutant splice site IVS20+5G→A (b and d) and normal splicing at mutant splice sites IVS23+10T→G and IVS38−10T→C (c and e). Clone IVS40+5G→A reveals both normal and abnormal splicing products, suggesting partial activity at the mutant site (a and f). The relative positions of the forward (⇀)and reverse (↼) RT-PCR primers, mutant sites (*), and the cryptic splice site in the pSPL3b vector sequence (○) are indicated. For sizing of RT-PCR products, the 100-bp ladder is given, and spans range 100–500 bp.