Investigation of crystallin genes in familial cataract, and report of two disease associated mutations

Abstract

Aims: Mutations of seven crystallin genes have been shown to cause familial cataract. The authors aimed to identify disease causing crystallin mutations in paediatric cataract families from south eastern Australia.

Methods: 38 families with autosomal dominant or recessive paediatric cataract were examined. Three large families were studied by linkage analysis. Candidate genes at regions providing significant LOD scores were sequenced. Single stranded conformational polymorphism (SSCP) analysis was used to screen five crystallin genes in the probands, followed by direct sequencing of observed electrophoretic shifts. Mutations predicted to affect the coding sequence were subsequently investigated in the entire pedigree.

Results: A LOD score of 3.72 was obtained at the gamma-crystallin locus in one pedigree. Sequencing revealed a P23T mutation of CRYGD, found to segregate with disease. A splice site mutation at the first base of intron 3 of the CRYBA1/A3 gene segregating with disease was identified by SSCP in another large family. Five polymorphisms were also detected.

Conclusions: Although mutations in the five crystallin genes comprehensively screened in this study account for 38% of paediatric cataract mutations in the literature, only two causative mutations were detected in 38 pedigrees, suggesting that crystallin mutations are a relatively rare cause of the cataract phenotype in this population.

Figure 1

Pedigree of ctas17, indicating the presence of the CRYGD P23T mutation. Shaded symbols indicate the presence of ophthalmologist confirmed cataract, and the proband is indicated by an arrow. + indicates a heterozygote; − indicates wild type; individuals without a + or − symbol were unavailable for genotyping.

Figure 2

(A) Restriction fragment length polymorphism analysis of the mutation at the donor splice site of exon 3 of the CRYBA1/A3 gene in pedigree crch08. Sizes of fragments in the molecular standard lane are indicated in base pairs (bp). Shaded symbols indicate the presence of ophthalmologist confirmed cataract, and the proband is indicated by an arrow. Wild type individuals display only the 263 bp band. Individuals heterozygous for the mutation display the undigested 263 bp band, as well as 172 bp and 91 bp cleavage products, although the latter band is not clear on the gel. (B) (i) Direct and (ii) retroilluminated pictures of sutural, nuclear, and peripheral cortical opacity typical of an affected patient in this pedigree (III:9). There is considerable intrapedigree variablility of the relative predominance of each component.