Association of a Branch–Site Consensus Sequence Polymorphism in the Pigment Epithelium–Derived Factor Gene within a Population with Age–Related Macular Degeneration

Abstract

Abstract: : Purpose: Pigment epithelium–derived factor (PEDF) is a retinal neurotrophic factor with antiangiogenic properties. Intravitreal levels of PEDF in patients with Age–Related Macular Degeneration (AMD) and Diabetic Retinopathy are known to be lower when compared to patients without proliferative disease. Efficient splicing of introns from pre–mRNA requires recognition of the 5’–donor splice site, branch–site and 3’–acceptor splice site sequences. Genetic alteration of the branch–site consensus sequence can cause human disease. We have identified a SNP within a branch–site consensus sequence of the PEDF gene. The purpose of this study is to determine the frequency of this branch–site SNP between patients with and without AMD and test for association of susceptibility to disease. Methods: The 5’ flanking promoter region including exon 1 and part of intron 1 of the PEDF gene from 206 patients with AMD (with neovascular AMD or geographic atrophy) and 176 controls were amplified and sequenced. All SNPs were scored and the frequency of each polymorphism was determined. Chi–squared and logistic regression analysis was performed to test for association with disease. Results: An adenosine (A allele) to a guanosine (G allele) polymorphism within a branch–site consensus sequence was identified. These alleles are in Hardy Weinberg equilibrium in cases and controls. Chi–squared test for homogeneity between AMD group and controls indicates that AMD affected individuals were significantly more likely (p=0.030) to carry at least one A allele. Logistic regression analysis shows that the presence of the A allele is significantly associated with AMD for women, p= 0.04; OR = 1.84; 95% CI (1.03, 3.28) in this population. A model including age as a predictor shows that both age and the presence of the A allele are significantly associated: Age p=0.0001; OR=0.40; 95% CI=0.25, 0.64 and A allele p=0.032; OR=1.94; 95% CI=1.06, 3.57. Conclusions: The SNP situated within the branch–site consensus sequence is at a position that when altered can abolish splicing. Further analysis is required to determine whether this branch–site polymorphism disrupts the normal splicing of intron 1. Linkage disequilibrium and haplotype analysis across this gene will further dissect the role of this region of the genome in susceptibility to AMD.

Keywords: age–related macular degeneration • genetics • candidate gene analysis