Evaluation of the CYP1B1 gene as a candidate gene in beagles with primary open-angle glaucoma (POAG)

Purpose: In humans, primary open-angle glaucoma (POAG) is a complex genetic disorder and is the leading cause of visual impairment. Although all relevant genes were not identified, a small subset of the condition is found to be caused by mutations in the MYOC and CYP1B1 genes. Inherited glaucoma also occurs in several breeds of dogs including beagles. Primary glaucoma in beagles is inherited as an autosomal recessive trait. The purpose of this study is to investigate the role of the CYP1B1 gene in beagles with POAG. Methods: For the purpose of genetic analysis, total RNAs from the spleen of the canines were isolated and CYP1B1 cDNA was prepared. Genomic DNA from five affected, two carriers, and 13 randomly selected normal beagles with no sign of glaucoma was amplified by the polymerase chain reaction (PCR) using four pairs of primers. The amplified products were directly sequenced using BigDye terminator cycle sequencing. Results: Genomic DNA analyses have identified a substitution polymorphism (109A -> C) in the 5'-untranslated region (UTR) as well as a missense mutation (P93R) in exon 2 of the gene. Three affected, two carriers, and nine normal dogs are heterozygous while two affected and three normal dogs are homozygous for the missense mutation. One normal dog did not show this alteration. Normal dogs also contain the substitution polymorphism in the 5'-UTR. Similar experiments with exon 3 did not identify any additional mutation in the gene. Conclusions: The above results suggest that CYP1B1 alterations in the coding and UTR are not the primary cause of glaucoma in beagles by possible monogenic association. They may be classified as polymorphisms or they may modify glaucoma phenotype.