Cellular cholesterol movement in cardiovascular disease

心血管疾病中的细胞胆固醇运动

• 批准号: 10397415
• 负责人: PETER J TONTONOZ
• 金额: $ 67.08万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-05-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10397415
• 关键词: Adrenal Cortex; Adrenal Glands; Area; Arterial Fatty Streak; Atherosclerosis; Binding; Binding Proteins; Biochemical; Biological; Biology; Biotin; Bone Marrow Transplantation; Cardiovascular Diseases; Cell membrane; Cells; Chemicals; Chemistry; Chimeric Proteins; Cholesterol; Cholesterol Esters; Collaborations; Data; Development; Disease; Electrons; Endoplasmic Reticulum; Esterification; Foam Cells; Functional disorder; Genes; Gonadal structure; HDL receptor; Health; High Density Lipoproteins; Homeostasis; Image; Impairment; In Vitro; Label; Laboratories; Lesion; Lipids; Mammalian Cell; Mass Spectrum Analysis; Measures; Mediator of activation protein; Membrane; Membrane Proteins; Metabolic Diseases; Metabolism; Molecular; Movement; Mus; Pathway interactions; Physiology; Proteins; Recording of previous events; Research Personnel; Role; SR-BI receptor; Series; Signal Pathway; Stable Isotope Labeling; Sterols; Testing; Therapeutic Intervention; Tissues; atherogenesis; chemoproteomics; cholesterol control; crosslink; defined contribution; imaging modality; in vivo; lipid metabolism; live cell microscopy; macrophage; member; mimetics; novel; novel strategies; particle; response; uptake

Project 3: Sterol Transport Pathways in Cardiovascular Disease ABSTRACT The objective of Project 3 is to define fundamental mechanisms that regulate cellular lipid flux and to elucidate their impact on systemic metabolism. Dissecting signaling pathways that govern how cells store, transport, and metabolize lipids is expected to uncover new opportunities for therapeutic intervention in metabolic disease. Although nonvesicular cholesterol transport has long been hypothesized to be critical for lipid homeostasis in mammalian cells, the underlying mechanisms have remained obscure. We have discovered a novel transporter called Aster-B that appears to fill this important gap in our understanding of sterol transport. Aster-B is a previously uncharacterized protein that facilitates the direct transport of cholesterol from the plasma membrane (PM) to the ER. We propose a series of molecular, cell biological, and mouse studies to investigate the roles of the Aster-B cholesterol transport pathway in physiology and disease. Aim 1 is to elucidate the role of Aster-B in cellular cholesterol transport, efflux, and esterification. We identified Asterb as a novel cholesterol-responsive LXR target gene. Gain or loss of Aster-B alters cholesterol distribution and impairs cholesterol ester synthesis in response to cholesterol loading. Using biochemical approaches and complementary imaging modalities including electron and live-cell microscopy, we will define the mechanism of action of Aster-B and its role in macrophage sterol flux. Aim 2 is to determine the impact of Aster-B on sterol transport in vivo. Preliminary data indicate that Asterb is most highly expressed in macrophages, adrenal gland, and gonads. We will determine the effect of loss of Aster-B expression on whole-body and tissue-specific lipid homeostasis. We hypothesize that Aster-B is a critical mediator of cellular cholesterol transport downstream of the HDL receptor SR-BI. Aim 3 is to define the contribution of the macrophage Aster pathway to atherosclerosis. The LXR pathway is one of the strongest known determinants of atherosclerotic lesion development. Our observation that Aster-B expression is regulated by LXRs suggests that Aster-dependent cholesterol transport may impact macrophage foam cell formation and the development of atherosclerosis. We will test the impact of gain or loss of Aster function on macrophage cholesterol uptake and efflux. We will perform bone marrow transplant studies into LDLR-deficient mice to test the impact of Aster-B deficiency on lesion formation. Aim 4 is to identify additional components of the Aster pathway. We will perform protein-interaction screens using a biotin proximity labeling strategy. We will perform a chemoproteomic screen using HDL particles loaded with a cholesterol-mimetic probe that can be crosslinked to proteins and retrieved using a click-chemistry handle. This application leverages the unique and complementary strengths of each member of our PPG, bringing together a range of approaches and technological capabilities that would be unavailable to any single investigator. Understanding the molecular pathways that control cholesterol movement in macrophages is central to the overall theme of this PPG application and will advance our understanding of both physiology and pathophysiology.

项目3：心血管疾病中的固醇运输途径 抽象的 项目3的目的是定义调节细胞脂质通量并阐明的基本机制 它们对系统性代谢的影响。解剖控制细胞如何存储，运输和 预计代谢脂质将发现用于代谢疾病的治疗干预的新机会。 尽管长期以来一直认为非抗胆固醇的转运对于脂质稳态至关重要 哺乳动物细胞，潜在的机制仍然晦涩难懂。我们发现了一个新颖的运输车 称为Aster-B，似乎填补了我们对固醇运输的理解。 Aster-B是一个 以前未表征的蛋白质促进胆固醇直接从质膜转运 （PM）到ER。我们提出了一系列分子，细胞生物学和小鼠研究，以研究 生理和疾病中的Aster-B胆固醇传输途径。目标1是阐明Aster-B在 细胞胆固醇转运，外排和酯化。我们将Asterb确定为一种新型胆固醇响应性 LXR靶基因。 Aster-B的增益或丧失会改变胆固醇的分布并损害胆固醇酯的合成 对胆固醇负荷的反应。使用生化方法和互补成像方式 包括电子和活细胞显微镜，我们将定义Aster-B的作用机理及其在 巨噬细胞固醇通量。 AIM 2是确定Aster-B对体内固醇转运的影响。初步数据 表明Asterb在巨噬细胞，肾上腺和性腺中最高表达。我们将确定 Aster-B表达丧失对全身和组织特异性脂质稳态的影响。我们假设 该Aster-B是HDL受体SR-BI下游细胞胆固醇转运的关键介质。目标3 是为了定义巨噬细胞陆上途径对动脉粥样硬化的贡献。 LXR途径是 动脉粥样硬化病变发育的最强决定因素。我们观察到Aster-B表达 受LXR的调节表明，依赖星号的胆固醇转运可能会影响巨噬细胞泡沫细胞 形成和动脉粥样硬化的发展。我们将测试Aster功能的增益或丧失对 巨噬细胞胆固醇吸收和外排。我们将对LDLR缺陷进行骨髓移植研究 小鼠测试Aster-B缺乏对病变形成的影响。目标4是确定的其他组件 Aster Pathway。我们将使用生物素接近标记策略执行蛋白质相互作用筛选。我们将 使用装有胆固醇模拟探针的HDL颗粒进行化学蛋白质筛选 与蛋白质交联，并使用点击化学手柄检索。此应用程序利用唯一的 我们PPG的每个成员的互补优势，汇集了一系列方法和 任何单个研究者都无法获得的技术能力。了解分子 控制巨噬细胞中胆固醇运动的途径对于此PPG的整体主题至关重要 应用并将提高我们对生理学和病理生理学的理解。