Cis-regulatory architecture of coronary vascular wall loci

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PROJECT SUMMARY Coronary artery disease (CAD) remains the leading cause of mortality worldwide and poses considerable public health burden. Both genetic and environmental risk factors contribute to CAD susceptibility, leading to increased disease prevalence in developing countries. Genome-wide association studies (GWAS) have now identified 161 independent genetic loci associated with CAD risk in large-scale meta-analyses in over 300,000 individuals. However, given that the majority of the associations reside in non-coding genomic regions, it has been challenging to translate these discoveries into biologically and clinically relevant insights. Interestingly, many of the candidate genes at CAD loci are organized into novel or unknown biological pathways, and point to dysregulation of vascular wall processes. Large efforts such as the Genotype Tissue Expression (GTEx) project and Stockholm-Tartu Atherosclerosis Reverse Network Engineering Task (STARNET) study have refined candidate gene regulatory mechanisms, however the specific cellular and phenotypic states driving these changes remains unclear. Smooth muscle cells (SMC) normally regulate vascular tone in the vessel wall but play critical roles in disease pathogenesis as their contractile gene program is hijacked during phenotypic switching to macrophage-like and fibroblast-like cells. Using epigenomic and expression quantitative trait loci (eQTL) mapping in 52 primary human coronary artery smooth muscle cells (HCASMC) we recently identified 11 candidate causal genes and mechanisms linking vascular wall processes to CAD risk (e.g. TCF21, SMAD3, CDKN2B, LMOD1). It is likely that non-coding regulatory variants function through changes in chromatin accessibility, which coincides with transcription factor (TF) binding in cis. By identifying these upstream TFs at vascular wall loci we can begin to assemble targetable pathways to modulate hidden disease risk in the vessel wall. In this proposal we plan to perform joint gene expression and chromatin accessibility QTL mapping in a unique cohort of 120 normal and diseased coronary artery tissues from explanted heart donors. We will validate and discover functional regulatory elements at GWAS loci using CRISPR/dCas9 perturbation with high-throughput phenotyping in SMC. Finally, we will investigate the cis-regulatory architecture in these samples at single-cell resolution and infer TF binding during phenotypic modulation. Together these studies will reveal the causal regulatory mechanisms responsible for CAD in the vascular wall and inform next generation treatment or prevention strategies to eradicate this debilitating disease.