Abstract Body: Introduction: Coronary artery disease (CAD) remains the leading cause of global mortality, with genetic factors conferring about 50% of disease susceptibility. GWAS have identified numerous genomic loci associated with CAD. However, translating these findings to clinical applications is largely impeded, due to significant gaps in our understanding of the underlying mechanisms, including sequence conservation, target genes(s), relevant cell types and biological functions. Vascular smooth muscle cells (VSMCs) contribute to CAD by transitioning from a contractile to a dedifferentiated state in response to stimuli. Therefore, identifying CAD-associated genes that regulate VSMC function is essential for understanding CAD etiology and improving therapeutic strategies.
Hypothesis: Novel CAD genes that primarily regulate VSMC function remain to be discovered.
Methods and Results: A total of 1,119 CAD-associated SNPs were collected from GWAS Catalog database (P<5e-08). Most of these risk SNPs are located within non-coding regions with potential regulatory function, together with 57 missense variants and 3 nonsense mutations. Sequence conservation analysis using PhastCons scores and cross-species comparation identified 246 SNPs that are conserved in mice, including 39 missense mutations. eQTL data from the GTEx database revealed 47.8% of non-coding SNPs associated with gene expression in aorta or coronary arteries. Integration of SNPs with scATAC-seq data from human atherosclerotic arteries demonstrated enrichment of CAD SNPs in SMC-specific open chromatin regions, which is consistent with previous reports and supports a SMC-specific regulatory mechanism. Analysis of scRNA-seq data further identified a list of SMC-enriched CAD genes. Notably, TNS2 and BDNF, harboring missense mutations, exhibited enriched expression in SMCs and phenotypically modulated SMCs, respectively. Additionally, SHROOM3 and IRAG1 gene, containing non-coding risk SNPs, were enriched in contractile SMCs and significantly downregulated in phenotypically modulated SMCs in atherosclerosis. The function of SHROOM3 and IRAG1 were then assessed by gene silencing in cultured human coronary artery SMCs.
Conclusion: This integrative analysis revealed novel CAD risk genes involved in VSMC function. The conserved SNPs offer opportunities for functional studies in mouse models. SHROOM3 and IRAG1 emerge as promising candidates for targeting VSMC function in CAD development.