9p21 DNA variants associated with Coronary Artery Disease impair IFNγ signaling response

Genome wide association studies (GWAS) have identified SNPs in the 9p21 gene desert associated with coronary artery disease (CAD)1–4 and Type 2 diabetes (T2D)5–7. Despite evidence for a role of the associated interval in neighboring gene regulation8–10, the biological underpinnings of these genetic associations to CAD or T2D have not yet been explained. Here we identify 33 enhancers in 9p21; the interval is the second densest gene-desert for predicted enhancers and 6 times denser than the whole genome (p<6.55 10(−33)). The CAD risk alleles of SNPs rs10811656/rs10757278 are located in one of these enhancers and disrupt a binding site for STAT1. Lymphoblastoid cell lines (LCL) homozygous for the CAD risk haplotype exhibit no binding of STAT1, and in LCL homozygous for the CAD non-risk haplotype binding of STAT1 inhibits CDKN2BAS expression, which is reversed by siRNA knock-down of STAT1. Using a new, open-ended approach to detect long-distance interactions (3D-DSL), we find that in human vascular endothelium cells (HUVEC) the enhancer interval containing the CAD locus physically interacts with the CDKN2A/B locus, the MTAP gene and an interval downstream of INFA21. In HUVEC, IFNγ activation strongly affects the structure of the chromatin and the transcriptional regulation in the 9p21 locus, including STAT1 binding, long-range enhancer interactions and altered expression of neighboring genes. Our findings establish a link between CAD genetic susceptibility and the response to inflammatory signaling in a vascular cell type and thus demonstrate the utility of GWAS findings to direct studies to novel genomic loci and biological processes important for disease etiology.