Role of Rare Human Genetic Variants in LRP1 on Aortic Aneurysm Formation

LRP1 中罕见的人类遗传变异对主动脉瘤形成的作用

• 批准号: 10192812
• 负责人: Allison Lindsay Arai
• 金额: $ 5.1万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10192812
• 关键词: 10 year old; Abdominal Aortic Aneurysm; Affect; Age; Age-Months; Alleles; Aneurysm; Aorta; Aortic Aneurysm; Aortic Rupture; Atherosclerosis; Attenuated; Binding; Biochemical; Bioinformatics; Blood Vessels; Cell surface; Cells; Childhood; Collaborations; Data; Defect; Development; Disease; Elderly; Embryo; Endoplasmic Reticulum; Exhibits; Failure; Genes; Genetic; Genetic study; Goals; Hemorrhage; Heterozygote; Homeostasis; Human; Human Genetics; Impairment; In Vitro; Inherited; LDL-Receptor Related Protein 1; Laboratories; Lead; Life; Ligand Binding; Ligands; Link; Low Density Lipoprotein Receptor; Macroglobulins; Mesoderm; Microscopy; Molecular Chaperones; Mothers; Mus; Mutation; Parents; Pathway Analysis; Pathway interactions; Patients; Pericytes; Pharmacologic Substance; Pharmacology; Phenotype; Physiological; Platelet-Derived Growth Factor Receptor; Population; Property; Proteins; Proteomics; Receptor Signaling; Recycling; Role; Rupture; Signal Pathway; Signal Transduction; Smooth Muscle Myocytes; Structure; Surface Plasmon Resonance; Testing; Thoracic Aortic Aneurysm; Time; Transforming Growth Factors; Up-Regulation; Vascular Smooth Muscle; age group; alpha 2-Glucoproteins; base; de novo mutation; exome sequencing; experimental study; genetic variant; improved; insight; loss of function; male; medical schools; mutant; new therapeutic target; novel; receptor; receptor binding; recruit; trafficking; uptake; western diet

PROJECT SUMMARY Abdominal aortic aneurysms (AAA) affect a significant portion of the population and are typically associated with advanced age and atherosclerosis. Until now, no single gene has been identified as causative for AAA. Our laboratory has initiated studies to characterize biochemical defects in a patient with severe presentation of AAA in childhood. Two mutations on separate alleles were identified in the low density lipoprotein (LDL) receptor- related protein 1 (LRP1) gene of the patient. The LRP1 mutations include a de novo mutation in a ligand binding region and an inherited mutation in a β-propeller domain, a structure that is critical for LRP1 trafficking to the cell surface. We predict that the functional deficits caused by each mutation are related to the physiological role of the structural component where each mutation occurs. The LRP1 gene encodes a large endocytic and signaling receptor that binds and traffics numerous ligands to lysosomal compartment for degradation. Mouse studies reveal that selective deletion of LRP1 in vascular smooth muscle cells (smLRP1-/-) results in spontaneous and fully penetrant aortic aneurysm formation. A major goal of these proposed studies is biochemical characterization of the functional defects imposed by these mutant receptors, which will be critical to defining mechanisms by which LRP1 regulates vessel wall homeostasis. Interestingly, bioinformatic pathway analysis of quantitative proteomic results revealed excess signaling of the transforming growth factor- β (TGF-β) pathway in both patient aortic smooth muscle cells and the aorta of smLRP1-/- mice compared to controls. I hypothesize that loss of LRP1 function in the patient smooth muscle cells results from impaired ligand binding due to the de novo LRP1 mutation and from inefficient delivery of LRP1 to the cell surface due to the β-propeller domain LRP1 mutation. Furthermore, I hypothesize that the impact of these LRP1 mutations results in a loss of ability to attenuate the TGF-β signaling pathway. This hypothesis will be tested in the following specific aims: (1) to define the mechanism by which mutations in LRP1 result in excessive TGF-β signaling and (2) to determine the impact of patient LRP1 mutations on LRP1 trafficking. In vitro experiments will determine if patient-specific LRP1 mutations alter TGF-β protein levels and signaling. Surface plasmon resonance experiments will test if patient-specific LRP1 mutations result in defective binding of LRP1 to TGF-β, TGF-β binding partners, or molecular chaperones involved in LRP1 trafficking. Proteomic experiments will define the LRP1 interactome in control and patient-derived aortic smooth muscle cells to further identify defects in mutant LRP1 molecules. Finally, microscopy-based experiments will examine trafficking and recycling properties of mutant forms of LRP1. This study will demonstrate the link between AAA and two independent LRP1 mutations. The results will improve understanding of AAA formation to develop better treatments and identify novel pharmaceutical targets.

项目总结 腹主动脉瘤(AAA)影响很大一部分人群，通常与 高龄和动脉粥样硬化。到目前为止，还没有单一的基因被确定为AAA的致病因素。我们的 实验室已开始研究一名严重腹主动脉瘤患者的生化缺陷。 在童年时。在低密度脂蛋白(LDL)受体上发现了两个独立的等位基因突变- 患者的相关蛋白1(LRP1)基因。LRP1突变包括配体结合中的从头突变 区域和β-螺旋体结构域中的遗传突变，这是LRP1向细胞转运的关键结构 浮出水面。我们预测，每个突变引起的功能缺陷都与其生理作用有关。 每一种突变发生的结构成分。LRP1基因编码一个大的内吞和信号 结合和运输大量配体到溶酶体隔间进行降解的受体。小鼠研究 揭示血管平滑肌细胞中LRP1的选择性缺失(smLRP1-/-)导致自发性和 完全穿透性主动脉瘤形成。这些拟议研究的一个主要目标是生化。 这些突变受体造成的功能缺陷的特征，这将是关键 确定LRP1调节血管壁内稳态的机制。有趣的是，生物信息学 定量蛋白质组学结果的通径分析显示，转化生长因子- β(转化生长因子-β)通路在患者和smLRP1-/-小鼠主动脉中的比较 控制。我假设患者平滑肌细胞中LRP1功能的丧失是由于 由于从头LRP1突变和LRP1向细胞表面的低效递送而导致的配体结合 这是由于β-螺旋体结构域LRP1突变所致。此外，我假设这些LRP1的影响 突变导致丧失减弱转化生长因子-β信号通路的能力。这一假设将是 测试的具体目标如下：(1)确定LRP1基因突变导致过量 转化生长因子-β信号转导和(2)确定患者LRP1突变对LRP1转运的影响。离体 实验将确定患者特有的LRP1突变是否会改变转化生长因子-β蛋白水平和信号转导。表面 等离子共振实验将测试患者特有的LRP1突变是否导致LRP1结合缺陷 与转化生长因子-β、转化生长因子-β结合伴侣或参与LRP1转运的分子伴侣有关。蛋白质组学实验 将在对照和患者来源的主动脉平滑肌细胞中定义LRP1相互作用组以进一步鉴定 突变的LRP1分子中的缺陷。最后，以显微镜为基础的实验将检查贩运和回收 LRP1基因突变形式的性质。这项研究将证明AAA和两个独立的 LRP1突变。这一结果将提高对AAA形成的理解，以开发更好的治疗方法和 确定新的药物靶点。