Abdominal aortic aneurysm is associated with a variant in low-density lipoprotein receptor-related protein 1

Abstract

Abdominal aortic aneurysm (AAA) is a common cause of morbidity and mortality and has a significant heritability. We carried out a genome-wide association discovery study of 1866 patients with AAA and 5435 controls and replication of promising signals (lead SNP with a p value < 1 × 10(-5)) in 2871 additional cases and 32,687 controls and performed further follow-up in 1491 AAA and 11,060 controls. In the discovery study, nine loci demonstrated association with AAA (p < 1 × 10(-5)). In the replication sample, the lead SNP at one of these loci, rs1466535, located within intron 1 of low-density-lipoprotein receptor-related protein 1 (LRP1) demonstrated significant association (p = 0.0042). We confirmed the association of rs1466535 and AAA in our follow-up study (p = 0.035). In a combined analysis (6228 AAA and 49182 controls), rs1466535 had a consistent effect size and direction in all sample sets (combined p = 4.52 × 10(-10), odds ratio 1.15 [1.10-1.21]). No associations were seen for either rs1466535 or the 12q13.3 locus in independent association studies of coronary artery disease, blood pressure, diabetes, or hyperlipidaemia, suggesting that this locus is specific to AAA. Gene-expression studies demonstrated a trend toward increased LRP1 expression for the rs1466535 CC genotype in arterial tissues; there was a significant (p = 0.029) 1.19-fold (1.04-1.36) increase in LRP1 expression in CC homozygotes compared to TT homozygotes in aortic adventitia. Functional studies demonstrated that rs1466535 might alter a SREBP-1 binding site and influence enhancer activity at the locus. In conclusion, this study has identified a biologically plausible genetic variant associated specifically with AAA, and we suggest that this variant has a possible functional role in LRP1 expression.

Figure 1

Whole-Genome Manhattan Plot of the Results of the Discovery Study p values expressed as a negative logarithm are plotted on the vertical axis and chromosomal positions of each SNP are plotted on the horizontal axis. SNPs with p values < 1 × 10⁻⁵ are shown in red, and represent the loci taken forward into the replication study.

Figure 2

GWAS Discovery Study Associations for the Region around rs1466535 Showing All Genes in the Region rs1466535 is indicated by the purple circle, the other SNPs typed in the discovery study are shown as filled circles with the color corresponding to their linkage with rs1466535. The solid blue line shows the regional recombination rate (right axis). Below the regional association plot is the LD plot for the same region with the standard D'/LOD Haploview color scheme. rs1466535 is highlighted by green lines.

Figure 3

Electrophoretic Mobility Shift Assay Demonstrating Allele-Specific Transcription Factor Binding to rs1466535 Lane A: binding of nuclear extract to labeled oligonucleotide containing rs1466535 T allele; lane B: T-allele binding with addition of excess unlabeled T-allele DNA competitor. The remaining band indicates a nonspecific protein-DNA interaction. Lane C: binding of nuclear extract to labeled oligonucleotide containing C allele. Lane D: C-allele binding with addition of excess unlabeled C-allele probe. Elimination of bands present in lane C indicates the binding of a specific protein. Lane E: C-allele-labeled oligonucleotide with addition of excess unlabeled fSREBP-1 (a transcription factor known to play an important role in LRP1 transcription) DNA competitor. The reduction in intensity of the lower band indicates that this binding is likely to be due to SREBP-1. Lane F: C-allele-labeled oligonucleotide with addition of excess non-SREBP-1-binding oligonucleotide competitor (sequence available on request) as a negative control. Lane G: binding of nuclear extract to labeled SREBP-1 consensus sequence, the lower band migrates with the putative SREBP-1 bound to the C allele. Lane H: SREBP-1 consensus sequence with the addition of excess SREBP-1 unlabeled competitor DNA. The lower band is eliminated, indicating this band represents the DNA-SREBP-1 complex. Together, these results suggest that rs1466535[T] removes a binding site for SREBP-1 in vitro.