A Role For Twist1 As Gwas Risk Gene For Coronary Artery Disease

Genome-wide association studies (GWAS) have identified rs2107595 as association SNP for both stroke and cardiovascular disease. This polymorphism is located proximal to the HDAC9 gene, with TWIST1 106kb downstream as the next closest gene in the recombination interval. Whilst studies in hyperlipidemic Hdac9 knockout mice have demonstrated a pro-atherogenic effect of Hdac9, recent evidence suggests an additionally role of TWIST1 in atherosclerosis. For one, rs2107595 genotypes associate with differences in TWIST1 but not HDAC9 expression levels in human aorta (GTEx consortium, n=197, p=0.00016), proposing a role of TWIST1 downstream the association locus specifically in vascular smooth muscle cells. RNA-seq data from in vitro cultured human coronary artery smooth muscle (HCASMCs) and endothelial cells (HCAECs) shows differential expression of TWIST1 in smooth muscle cells (n=19/20, q=3E-60). Furthermore, TWIST1 expression is selectively upregulated in HCASMCs upon serum starvation, and TWIST1 protein is detected in human lesions both near the tunica media and at the fibrous cap where it co-localizes with smooth muscle actin (ACTA2). The GWAS association SNP is predicted to change transcription factor binding affinity from the more general E2F to RBPJ - a well-known modulator of Notch signaling in smooth muscle cells (Transfac professional, 2014.4 data release). The variant is located within an enhancer region in mesenchymal stem cells (Broad GSE17312), fibroblasts (UCSF-UBC-USD GSE16368) as well as in HCASMCs. Current studies explore the tissue-specific activity of this regulatory element through in vivo lacZ reporter assays to investigate a potential role in mouse coronary arteries. Genome editing studies in immortalized HCASMCs are being employed to investigate the effect of SNP genotype on expression levels of both TWIST1 and HDAC9. Future experiments using an inducible, smooth muscle specific knockout of Twist1 will interrogate the potential role of Twist1 in atherosclerosis.