Uncommon Rearrangement and De Novo Mutation of the RPE65 Gene Associated With Leber Congenital Amaurosis (LCA)

Abstract

Purpose: : To understand the uncommon segregation of RPE65 mutations in two unrelated LCA patients.

Methods: : Two unrelated patients whom clinical history strongly suggested the involvement of the RPE65 gene were screened for mutation using DHPLC and direct sequencing. Subsequently, haplotype analyses at the RPE65 locus on chromosome 1p31 were carried out in both families using polymorphic markers flanking RPE65. The paternity was checked for the two patients using numerous markers of chromosome 1 as well as 8 highly polymorphic markers localized on 4 distinct chromosomes.

Results: : Apparent homozygosity for an already reported RPE65 splice–site mutation (c.11+5G>A) was identified in one of the two patients. The segregation analysis of the mutation showed that the mother was heterozygote for this mutation while the father did not carry it. Subsequent haplotype studies demonstrated homozygosity for 6 markers flanking the mutation, showing no paternal contribution for a 31 Mb region lying between the 1p31.1 and 1p22.1 chromosomal bands. On either side of this homoallelic region, correct bi–parental transmission was noted for the whole chromosome 1. Besides, the paternity was confirmed using markers of four other chromosomes. The second patient was found to be compound heterozygous for the same splice–site mutation (c.11+5G>A) and a missense mutation lying in exon 5 (c.407T>G, p.Val136Gly). The familial segregation showed that the c.11+5G>A mutation was inherited from the mother but the missense was absent in both the father and the mother. Subsequent haplotype studies were performed using four markers flanking the mutation confirming the correct bi–parental transmission. In addition, the paternity was confirmed using markers of four other chromosomes.

Conclusions: : It is worth noting that a paternal isodisomy of the whole chromosome 1 had been reported previously in a patient homozygous for the splice–site mutation c.11+5G>A. In this study, we report the identification of two uncommon inheritance phenomenons associated with the same splice–site mutation. In the first case, the apparent 30 Mb homoallelic region suggests the possibility of a partial maternal isodisomy or a de novo large deletion. In the second case, the identification of a RPE65 mutation not inherited from his parents suggests the occurrence of a de novo mutation.

Keywords: retinal degenerations: hereditary • candidate gene analysis • mutations