The BCAR1 locus in carotid intima-media thickness and atherosclerosis

Carotid intima media thickness (IMT) is a marker of subclinical atherosclerosis that can predict cardiovascular events over traditional risk factors. This thesis focused on a chromosome 16 locus associated with IMT and coronary artery disease, in order to identify functional variation affecting protein structure or gene expression, and investigate the role of novel genes in atherosclerosis. Bioinformatics tools were used to identify variants in strong LD with the lead SNP and to filter these to a shortlist of potential regulatory variants. Electrophoretic mobility shift assays on these 6 SNPs detected allele-specific protein binding to the lead SNP rs4888378 in CFDP1, and implicated the protein FOXA. Luciferase reporter assays showed a 35-92% decrease in gene expression with the A allele. Expression-QTL analysis confirmed associations of the protective allele of rs4888378 with higher expression of BCAR1 in vascular tissues. Genotyping and analysis of the lead SNP in the PLIC cohort of 2144 healthy men and women suggested a sex-specific effect of the SNP on IMT progression. Meta-analysis of five cohort studies supported a protective effect of the A allele on common-carotid IMT in women only. As illustrated by this locus, functional variation often lies in enhancers far from its target promoter. Circular chromosome conformation capture (4C) was explored to investigate interactions between enhancers and promoters at this locus, with the aim of capturing regions interacting with rs4888378 and the BCAR1 promoter. Analysis of exome sequencing data identified a SNP in BCAR1 (coding amino acid change P76S), which was genotyped in two cohorts (IMPROVE and PLIC). Associations with lower plaque and IMT were found (wild-type proline form), but differed between the cohorts and require further validation. Wild-type and variant vectors were expressed in cells to assess the variant’s effect on protein function and pathways involved in plaque, showing cells with wild-type protein to migrate more quickly. These analyses on regulatory and protein-coding mechanisms implicate BCAR1 in atherosclerosis and provide new pathways for analysis in the understanding of cardiovascular disease