From genotype to phenotype in a GWAS locus: the role of REST in atherosclerosis

项目摘要 这个K 08指导临床科学家研究职业发展奖是一个为期五年的计划，旨在 促进Marios Arvanitis博士（PI）发展成为血管遗传学领域的独立医生-研究者。 动脉粥样硬化性心血管疾病（ASCVD）是一种主要的公共卫生负担， 美国每年有60万人死亡。ASCVD是高度遗传和全基因组关联研究 已经发现了许多增加人群中疾病风险的候选基因组位点，从而 为新疗法提供了一个窗口。然而，大多数ASCVD的基因组风险位点在 它们是如何导致疾病风险的。 PI先前发表的工作集中在基因组风险位点的机制解释上 包括开发一种新的贝叶斯方法，称为CAFEH， 靶组织和基因组位点中的基因。我们对ASCVD遗传基础的初步分析 揭示了内皮细胞富集ASCVD遗传性，我们已经使用这些方法来优先考虑 预测通过改变RE-1沉默的表达来影响ASCVD风险的4号染色体基因座 转录因子（REST）基因在内皮细胞。K 08项目将通过以下方式探索监管机制： REST位点和基因影响动脉粥样硬化的发展。Aim 1将使用CRISPR-Cas9 在人类干细胞中进行编辑，然后将其分化为内皮细胞，以识别致病变体 以及在4 q12冠状动脉疾病GWAS基因座中介导相关性的上游转录因子。 目的2将确定内皮细胞中受REST影响的远端基因和通路，并研究其细胞内表达。 结果，从评估REST在内皮向间充质转化中的作用开始。目标3将使用 他莫昔芬诱导的内皮特异性Rest基因敲除小鼠模型，以评估Rest在体内的作用 内皮和动脉粥样硬化。 该项目的成功得到了多学科指导团队的支持，包括 一位血管生物学家（查尔斯·洛温斯坦博士），一位计算生物学家（亚历克西斯·巴特尔博士）和一位功能生物学家。 遗传学专家安德鲁·麦卡连博士（Andrew McCallion），沿着血管生物学专家咨询委员会， 细胞分化和动脉粥样硬化（Harry Dietz博士、Chulan Kwon博士和托马斯Quertermous博士）。这 奖励期将帮助PI提高他们的基因组学技能，获得新的湿实验室技能并生成初步数据 成功竞争R 01资金，以便将遗传见解转化为新的机制， ASCVD。

Project Summary This K08 mentored clinical scientist research career development award is a five-year program designed to facilitate Dr. Marios Arvanitis’ (PI) development into an independent physician-investigator in vascular genetics. Atherosclerotic cardiovascular disease (ASCVD) is a major public health burden that accounts for over 600,000 deaths in the United States each year. ASCVD is highly heritable and genome-wide association studies have discovered many candidate genomic loci that increase the risk of the disease in the population, thereby providing a window to novel therapies. However, most genomic risk loci for ASCVD remain unexplored in terms of how they lead to disease risk. Previously published work by the PI has focused on the mechanistic interpretation of genomic risk loci for cardiovascular disease, including the development of a novel Bayesian method, called CAFEH, to prioritize the target tissue and genes in genomic loci. Our preliminary analyses of the genetic underpinnings of ASCVD reveal that endothelial cells are enriched for ASCVD heritability, and we have used those methods to prioritize a chromosome 4 locus that is predicted to affect ASCVD risk via altering the expression of the RE-1 silencing transcription factor (REST) gene in endothelial cells. This K08 project will explore the regulatory mechanisms via which the REST locus and gene influence the development of atherosclerosis. Aim 1 will employ CRISPR-Cas9 editing in human stem cells which will then be differentiated into endothelial cells to identify the causal variants and the upstream transcription factors that mediate the association in the 4q12 coronary disease GWAS locus. Aim 2 will define distal genes and pathways affected by REST in the endothelium and investigate their cellular consequences, starting with evaluating the role of REST in endothelial to mesenchymal transition. Aim 3 will use a tamoxifen inducible endothelial specific Rest knock-out mouse model to evaluate the in vivo effects of Rest in the endothelium and atherosclerosis. The success of this project is guaranteed by the support of a multidisciplinary mentoring team including a vascular biologist (Dr. Charles Lowenstein), a computational biologist (Dr. Alexis Battle), and a functional genetics expert (Dr. Andrew McCallion), along with an advisory committee of experts in vascular biology, stem cell differentiation and atherosclerosis (Dr. Harry Dietz, Dr. Chulan Kwon and Dr. Thomas Quertermous). This award period will help the PI boost their genomics skills, acquire new wet lab skills and generate preliminary data to successfully compete for R01 funding in order to translate the genetic insights into novel mechanisms for ASCVD.