PDGFD regulates a transcriptional network to modulate smooth muscle cell transition and coronary artery disease risk

PDGFD 调节转录网络以调节平滑肌细胞转变和冠状动脉疾病风险

基本信息

批准号：
10593934
负责人：
THOMAS QUERTERMOUS
金额：
$ 67.51万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-04-15 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10593934
关键词：
AHR gene ATAC-seq Alleles Anatomy Animals Apolipoprotein E Aryl Hydrocarbon Receptor Atherosclerosis Autocrine Communication Binding Blood Vessels CRISPR interference Cardiovascular Diseases Cell Lineage Cell model Cells Chromatin Chromosome Mapping Complement Factor D Coronary Arteriosclerosis Coronary artery Coupled Data Development Disease Enhancers Epigenetic Process Fibroblasts Functional disorder Gene Expression Gene Targeting Genes Genetic Risk Genetic Transcription Goals Growth Factor Human Human Genome In Vitro Knock-out Knockout Mice LacZ Genes Link MADH3 gene Maps Measures Mediating Modeling Molecular Morphology Mus PDGFA gene PDGFRB gene Phenotype Platelet-Derived Growth Factor Probability Process Regulation Reporter Research Risk Assessment Role Scientist Signal Pathway Signal Transduction Smooth Muscle Myocytes Stromal Cell-Derived Factor 1 Subcellular Anatomy TWIST1 gene Therapeutic Tissues Transcription Process Transcriptional Activation Wild Type Mouse Work cell dedifferentiation cell type disorder risk gene function genome wide association study genome-wide in vivo mouse model programs public health relevance receptor response single-cell RNA sequencing transcription factor transcriptomic profiling transcriptomics vascular stress whole genome

项目摘要

We have identified TCF21 as the coronary artery disease (CAD) associated gene mapped by genome-wide association studies at 6q23.2 and employed numerous mechanistic approaches to show that it promotes a smooth muscle cell (SMC) transition to a fibroblast like “fibromyocyte” phenotype, and the contribution of these cells to the protective fibrous cap. Our studies with another CAD associated gene, the aryl hydrocarbon receptor (AHR), have characterized the transition of SMC to a second, chondrogenic “chondromyocyte” phenotype. To extend this work and investigate the mechanisms of epigenetic signaling upstream of TCF21, AHR, and other factors that mediate SMC cell state, we are focusing efforts on the CAD associated platelet derived growth factor D gene (PDGFD). We have shown that PDGFD regulates TCF21 and other validated CAD genes including LMOD1, CXCL12, and SMAD3, and is expressed primarily in disease transition SMC that also express the PDGFRB receptor. Together, these data suggest that PDGFD activates an autocrine signaling pathway that modulates SMC phenotype and CAD risk. The hypothesis directing this research postulates 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. The primary goals of the work proposed here are thus to identify the PDGFD target transcription factors (TFs) that regulate SMC transitions and characterize their transcriptional program in this cell type. Specifically, in Aim 1 we will employ Pdgfd knockout and SMC lineage tracing in the ApoE null mouse atherosclerosis model to characterize the effect of this gene on SMC cell state transitions, and the impact of perturbing these transitions on disease morphology and cellular anatomy. In Aim 2, we will conduct single cell RNA sequencing (scRNAseq) in Pdgfd null and wildtype atherosclerotic mice to characterize the SMC gene expression program downstream of Pdgfd in this cell type. Single cell ATAC sequencing (scATACseq) in the same animals will map enhancers genome- wide that are differentially regulated in SMC phenotypic transitions, and identify specific TFs that bind these enhancers to regulate expression of fibromyocyte and chondromyocyte specific genes. In Aim 3, we will perturb candidate SMC transition promoting TFs that are identified in Aim 2, in vitro in a PDGFD stimulated human coronary artery smooth muscle cell de-differentiation model, and the resulting transcriptomic and cell state effects interpreted in the context of PDGFD function in this model. These studies will link PDGFD to CAD associated genes that we have characterized in the context of SMC phenotypic transition (TCF21, AHR, SMAD3, TWIST1), and to additional high probability CAD genes that regulate SMC phenotype, to expand the disease transcriptional network in this vascular cell type. This work will advance our understanding of atherosclerosis pathophysiology and promote efforts to target vascular wall molecular processes to ameliorate CAD risk.

我们已经确定 TCF21 是通过全基因组图谱定位的冠状动脉疾病 (CAD) 相关基因 6q23.2 的关联研究并采用了多种机制方法来表明它促进了平滑肌细胞 (SMC) 转变为类似“纤维肌细胞”表型的成纤维细胞，以及这些细胞的贡献我们对另一种 CAD 相关基因——芳基碳氢化合物的研究。受体（AHR），表征了 SMC 向第二种软骨形成“软骨肌细胞”的转变为了扩展这项工作并研究 TCF21 上游的表观遗传信号机制， AHR 以及介导 SMC 细胞状态的其他因素，我们将重点放在与 CAD 相关的血小板上衍生生长因子 D 基因 (PDGFD) 我们已经证明 PDGFD 调节 TCF21 和其他经过验证的基因。 CAD 基因包括 LMOD1、CXCL12 和 SMAD3，主要在疾病转变 SMC 中表达，也表达 PDGFRB 受体，这些数据表明 PDGFD 激活自分泌。调节 SMC 表型和 CAD 风险的信号通路。假设 PDGFD 通过调节 TCF21 和其他关键疾病来增加 CAD 风险介导 SMC 对血管应激表型反应的相关转录因子。因此，本文提出的工作目标是确定调节 PDGFD 的靶转录因子 (TF) 具体而言，我们将在目标 1 中进行 SMC 转变并表征其转录程序。在 ApoE 缺失小鼠动脉粥样硬化模型中采用 Pdgfd 敲除和 SMC 谱系追踪来表征该基因对 SMC 细胞状态转变的影响，以及干扰这些转变对疾病的影响在目标 2 中，我们将在 Pdgfd 中进行单细胞 RNA 测序 (scRNAseq)。无效和野生型动脉粥样硬化小鼠来表征 Pdgfd 下游的 SMC 基因表达程序在这种细胞类型中，对同一动物进行单细胞 ATAC 测序 (scATACseq) 将绘制增强子基因组图谱。广泛的 SMC 表型转变中受到差异性调节，并识别结合这些的特定 TF 在目标 3 中，我们将使用增强子来调节纤维肌细胞和软骨肌细胞特异性基因的表达。在PDGFD刺激的体外干扰目标2中鉴定的候选SMC转变促进TF 人冠状动脉平滑肌细胞去分化模型，以及由此产生的转录组和细胞在该模型中，在 PDGFD 功能的背景下解释状态效应，这些研究将 PDGFD 与 CAD 联系起来。我们在 SMC 表型转变的背景下表征的相关基因（TCF21、AHR、 SMAD3、TWIST1），以及调节 SMC 表型的其他高概率 CAD 基因，以扩展这项工作将增进我们对这种血管细胞类型的疾病转录网络的理解。动脉粥样硬化病理生理学并促进针对血管壁分子过程的努力以改善加元风险。