Abstract
Purpose: :
Glaucoma is a leading cause of blindness in the United States. Important risk factors contributing to glaucoma include elevated IOP, positive family history, age, race, and thin central corneal thickness (CCT). Our goal is to identify genes that regulate the magnitude of CCT, using it as an entry point for studying the etiology of glaucoma. Using a quantitative approach with intercrosses between mice with thin corneas (C57BLKS/J; KS) and thick corneas (SJL/J; SJL), we identified Central corneal thickness QTL 1 (Cctq1), which spans a ~15 cM region on mouse chromosome 7. The purpose of these experiments is to fine-map the position of Cctq1, identify known DNA variations within the Cctq1 interval using bioinformatics, and to begin testing candidate genes.
Methods: :
To map the Cctq1 gene, the genomic interval was divided into fourths (i.e., Intervals I, II, III, and IV) by intercrossing KS.Cctq1KS/SJL N4F2 congenic mice. Mice harboring recombinant chromosomes were identified by following genotypes at 5 polymorphic markers (D7Mit318, D7Mit31, D7Mit321, D7Mit220, and D7Mit238). These recombinant mice (and controls) were phenotyped for CCT, and differences in CCT were statistically analyzed using ANOVA. To sequence candidate genes (Pak1 and Aqp11), exons were PCR amplified and sequenced.
Results: :
Congenic mice harboring either Cctq1 interval I, II or III all confer a significant increase in CCT compared to control mice (P < 0.01). Mice harboring Cctq1 interval IV have CCT values not statistically different than control mice. Using bioinformatics, we identified 8 known nonsynonymous polymorphisms within the Cctq1 interval, the most interesting of which is within Adamtsl3. Of candidates sequenced, none of the exons contained any DNA variations between KS and SJL parental strains of mice.
Conclusions: :
Our results suggest at least two possibilities: 1) more than one CCT-regulating gene exists within Cctq1; or 2) there is too much genetic heterogeneity in our N4 mice to be able to uncover the specific allele. To differentiate between the two possibilities, we are continuing the backcross to N10 to reduce the genetic heterogeneity. We are also continuing candidate gene sequencing.
Keywords: gene mapping • cornea: basic science • genetics