We have identified TCF21 as the coronary artery disease (CAD) associated gene mapped by genome-wide association studies at 6q23.2. By combining conditional deletion of Tcf21, smooth muscle cell (SMC) lineage tracing, single cell RNA sequencing (scRNAseq), and anatomical cellular lesion analysis in the ApoE null model, we have shown that it 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. This work profiled at a single cell level the transition of SMC to a fibroblast like phenotype, creating cells that we term ?fibromyocytes? (FMC). Genomic studies conducted as part of this funded work have suggested that TCF21 binds and regulates expression of a number of cooperating transcription factors (TFs) in other CAD loci to govern the SMC-FMC transition. Further, TCF21 targeted TFs in other CAD loci modulate an SMC transition to a chondrocyte-like phenotype, which is characterized by gene expression patterns typical of endochondral bone formation, producing cells we term ?chondromyocytes? (CMC). These findings point to two interrelated complex gene networks that regulate SMC cell state transition as a mechanism of disease causality. Our hypothesis for this renewal application thus proposes that: disease risk associated with SMC phenotypic transition is mediated by TCF21 and related transcription factors that regulate interactive transcriptional networks constituted in large part by CAD associated genes. The primary goal of work proposed here is to further characterize these networks and define the epigenetic and transcriptional mechanisms of TF interactions that determine the CAD risk engendered by the SMC phenotypic response to vascular stress. Specifically, in Aim 1 we will conduct single cell ATAC sequencing (scATACseq) with wildtype and Tcf21 null atherosclerotic mice, as well as human coronary artery tissues, to map enhancers genome-wide that are differentially regulated in SMC phenotypic transitions, and identify TFs that bind these enhancers.
In Aim 2 we will perform scATACseq and scRNAseq following CRISPR/Cas9 perturbation of identified transition TFs in a human coronary artery smooth muscle cell de-differentiation model to examine the impact of TF knockdown on transcriptional profiles and interactions of TFs linked to the FMC and CMC phenotypes. To investigate the relationship of SMC phenotype to CAD risk, we will determine how perturbation of SMC transition TFs alters accessibility at promoter regions and linked enhancer regions at CAD associated loci.
In Aim 3 we will examine with molecular methods the mechanisms of epistasis and functional interactions between the TFs that primarily define the SMC transition phenotypes and identify transcriptional links to CAD. This work will thus characterize fundamental processes by which SMC TFs activate phenotypic transitions through epigenetic and transcriptional mechanisms, extending our understanding of this disease and promoting opportunities for ameliorating human CAD risk.
) Significant expense and effort by groups of scientists around the world has led to identification of regions of the human genome that are associated with the genetic risk for various forms of cardiovascular disease, including coronary artery disease. Additional research is required to understand the specific genes involved, and how they work to contribute to the disease process. Such information will allow for better risk assessment and the development of better therapeutics for these diseases.
| Liu, Boxiang; Pjanic, Milos; Wang, Ting et al. (2018) Genetic Regulatory Mechanisms of Smooth Muscle Cells Map to Coronary Artery Disease Risk Loci. Am J Hum Genet 103:377-388 |
| Paik, David T; Tian, Lei; Lee, Jaecheol et al. (2018) Large-Scale Single-Cell RNA-Seq Reveals Molecular Signatures of Heterogeneous Populations of Human Induced Pluripotent Stem Cell-Derived Endothelial Cells. Circ Res 123:443-450 |
| Carcamo-Orive, Ivan; Huang, Ngan F; Quertermous, Thomas et al. (2017) Induced Pluripotent Stem Cell-Derived Endothelial Cells in Insulin Resistance and Metabolic Syndrome. Arterioscler Thromb Vasc Biol 37:2038-2042 |
| Pjanic, Milos; Miller, Clint L; Wirka, Robert et al. (2016) Genetics and Genomics of Coronary Artery Disease. Curr Cardiol Rep 18:102 |
