Functional analysis of a carotid intima-media thickness locus implicates BCAR1 and suggests a causal variant

BACKGROUND: -Carotid intima-media thickness (cIMT) is a marker of subclinical atherosclerosis that can predict cardiovascular disease (CVD) events over traditional risk factors. This study examined the BCAR1-CFDP1-TMEM170A locus on chromosome 16, associated with cIMT and coronary artery disease in the IMPROVE cohort, to identify the functional variant. METHODS AND RESULTS: -In analysis of the locus's lead SNP (rs4888378, intronic in CFDP1) in Progressione della Lesione Intimale Carotidea (PLIC), the protective AA genotype was associated with slower IMT progression in women (p=0.04) but not men. Meta-analysis of five cohort studies also supported a protective effect of the A allele on common-carotid IMT in women only (women: β=-0.0047, p=1.63×10-4; men: β=-0.0029, p=0.0678). 214 non-coding variants in strong Linkage Disequilibrium (LD) (r2≥0.8) with rs4888378 were identified from 1000 Genome Project. ENCODE regulatory chromatin marks were used to create a shortlist of six possible regulatory variants. Electrophoretic mobility shift assays (EMSA) on the shortlist detected allele-specific protein binding to the lead SNP rs4888378; multiplexed-competitor EMSA implicated FOXA as the protein. Luciferase reporter assays on rs4888378 showed a significant 35-92% (p = 0.0057; p=4.0×10-22) decrease in gene expression with the A allele. Expression-QTL analysis confirmed previously reported associations of rs4888378 with BCAR1 in vascular tissues. CONCLUSIONS: -Molecular studies suggest the lead SNP as a potentially causal SNP at the BCAR1-CFDP1-TMEM170A locus, and eQTL studies implicate BCAR1 as the causal gene. This variant showed stronger effects on common-carotid IMT in women, raising questions about the mechanism of the causal SNP on atherosclerosis.