PROJECT SUMMARY OR ABSTRACT There are more than 160 genetic loci discovered which are associated with coronary artery disease (CAD), yet, outside of lipid lowering therapies, the promise of these genome wide association studies (GWAS) to identify causal genes and result in novel mechanisms for treatment has not been fulfilled. It is of critical need to identify causal genes and characterize the cellular signaling, transcriptomic, and epigenomic regulation of disease to discover novel treatments of CAD. The lab of my mentor has identified TCF21 as the CAD associated gene mapped by GWAS at 6q23.2. Single cell transcriptomic analysis and disease lesion cellular anatomy studies show that TCF21 is upregulated in SMC to promote de-differentiation, proliferation, and migration of medial SMC into the plaque where they contribute to the protective fibrous cap. We have begun to study the upstream regulation of TCF21 expression by the CAD associated gene encoding platelet derived growth factor D (PDGFD). PDGFD is a member of a receptor tyrosine kinase ligand family that has been well studied in the context of SMC phenotype and atherosclerosis and has a genetic association with CAD in humans. Gene variants resulting in increased PDGFD expression are associated with worsened CAD. These findings have led us to focus our attention on its determinative role in CAD. Importantly, PDGFD also directly regulates other validated CAD genes, including CDKN2B, LMOD1, CXCL12, SMAD3, and TWIST1, thus suggesting that it has a fundamental role in the modulation of CAD risk. We hypothesize that PDGFD promotes CAD risk through its regulation of TCF21 and other key disease related transcription factors that mediate the SMC phenotypic response to vascular stress. To address this hypothesis, we will use a novel Pdgfd-/- x ApoE- /- mouse model on a smooth muscle specific reporter background in combination with state-of-the-art technology to simultaneously evaluate single cell gene transcription and chromatin accessibility through combination scRNAseq with scATACseq in vascular tissue. We will address our primary hypothesis with two major specific aims, 1) Define the role of Pdgfd in regulating SMC phenotype and cellular and molecular features of atherosclerotic vascular tissues with an in-vivo mouse model, and 2) Map the transcriptomic program and causal epigenetic regulatory features that mediate the disease related cellular and molecular effects of Pdgfd in-vivo in the atherosclerosis mouse knockout model. This study will give us a fundamental understanding of how PDGFD influences SMC phenotypic response and thus CAD risk through modulation of chromatin accessibility and transcription factor binding. Completion of this proposal will address fundamental questions: (1) How does perturbation of Pdgfd expression affect the SMC phenotype and molecular features of atherosclerotic vascular tissue in vivo? (2) How does Pdgfd activate epigenetic and transcriptional processes which media...