Refinement of the Locus for X–Linked Congenital Cataract (CXN) and Positional Cloning of the Nance–Horan Syndrome (NHS) Gene

Abstract

Abstract: : Purpose: CXN was mapped to a 3 cM interval on chromosome Xp22, which is syntenic to the X–linked mouse cataract disease locus Xcat and encompasses the recently refined NHS locus. To identify the gene(s) responsible for these diseases, families with NHS were ascertained for molecular genetic analysis, we attempted to refine the disease interval for CXN, and candidate genes were analysed. Methods: Eight microsatellites were analysed within 21 individuals of the CXN family using the ABI 3100. Candidate genes were screened by PCR and direct sequencing of coding exons and intron–exon splice sites. Genomic structures were determined using comparative bioinformatics. Expression studies were performed using specific exonic primers to amplify commercial human cDNA, and extracted mouse RNA. Results: Haplotypes were reconstructed confirming disease segregation with markers on Xp22.13. A proximal cross over between novel markers S3 and S5 refined the CXN disease interval by approximately 300 Kb, flanked by markers DXS9902 and S5, excluding two genes PPEF1 and CDKL5 as candidates for disease. Two known (RBBP7 and RAI2) and two novel genes (from contig NT_011757) were screened for mutations in an affected male from the CXN family and a NHS family. Mutations in one of these genes were detected in two NHS families, both of which are predicted to cause protein truncations. Conclusions: We used the refined locus for CXN to focus the search for candidate genes. Mutations in this novel gene (NHS) were identified as causative for NHS. This gene was also independently cloned and recently reported as causative for NHS (Burdon et al. 2003). NHS encodes a protein of at least 1,630 aa and has no similarity to other proteins. We found expression of NHS in human fetal brain, thymus, kidney, adult retinal cDNA and mouse tooth cDNA at day 19. To date, no mutation has been identified in the CXN family and further characterisation of the genomic structure of NHS is underway.

Keywords: cataract • positional cloning • genetics