Intracellular Cholesterol Transport

细胞内胆固醇转运

• 批准号: 10059259
• 负责人: Frederick R. Maxfield
• 金额: $ 33.9万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-01-01 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10059259
• 关键词: Affect; Atherosclerosis; Autophagosome; Binding; Biological; Biological Assay; Bone Marrow; Cell Line; Cell membrane; Cell physiology; Cells; Cholesterol; Cholesterol Homeostasis; Complex; Computational Biology; Computer Analysis; Computer Models; Crystallization; Data; Development; Feedback; HepG2; Hepatocyte; Hereditary Disease; Human; In Vitro; Lipids; Liposomes; Maintenance; Mediating; Membrane; Methods; Modeling; Molecular; Mus; Mutation; NMR Spectroscopy; Organelles; Phosphatidylinositol Phosphates; Physiological; Play; Property; Protein Dynamics; Proteins; Protocols documentation; Recycling; Reporting; Role; Site; Specificity; Sterols; Structural Protein; Structure; Testing; Work; X-Ray Crystallography; cell type; cholesterol control; computer studies; experimental analysis; experimental study; knock-down; late endosome; macrophage; molecular dynamics; mutant; novel; structural biology; uptake; vector

Cholesterol plays an essential role in determining the properties of biological membranes. Control of cholesterol levels in organelles depends on sterol transport mechanisms that are poorly understood. Novel methods to study sterol transport among organelles quantitatively have been developed, and this is leading to precise models for transport among organelles. Preliminary data show that a widely expressed, sterol- regulated, soluble sterol transporter, STARD4, plays an important role in nonvesicular transport of cholesterol, and its role and mechanism of transport will be studied. STARD4 is an important component of a complex sterol regulatory network. Recent work showed that STARD4 is responsible for about 25% of sterol transport between two major pools – the plasma membrane and the endocytic recycling compartment - in U2OS cells. The role of STARD4 will also be assessed in macrophages, a cell type in which cholesterol metabolism plays a major role in development of atherosclerosis, and in HepG2 cells, a model for hepatocytes. In preliminary studies it has been found that STARD4 has unique interactions with phosphatidylinositol phosphates (PIPs). STARD4 extraction of sterol from membranes is accelerated nearly 10-fold when the membranes contain PI(4,5)P2, but there is no effect of PI(4,5)P2 in membranes accepting sterol from STARD4. Conversely, PI(3,5)P2 or PI5P in acceptor membranes accelerates transfer ∼10-fold with no effect when these PIPs are in donor membranes. PI3P has a similar but smaller effect. The role of STARD4 in sterol transport to late endosomes, lipid droplets, and autophagosomes (i.e., organelles with good acceptor PIPs) will be analyzed. Molecular dynamics simulations of interactions of STARD4 with membranes is identifying potential sites of interaction with PIPs. The effects of mutations of these sites will be assayed in a liposome to liposome sterol transport assay. Mutant STARD4 molecules with altered PIP sensitivity will be expressed in cells to determine effects on sterol transport and distribution. In preliminary studies, mutants identified by molecular dynamics were confirmed by tests in the liposome assay, and it was found that they do affect transport of sterol among organelles. X-ray crystallography has identified differences between human STARD4 with and without bound sterol. NMR spectroscopy is being used to analyze STARD4 protein dynamics and to determine the sites of interaction with lipids containing various PIPs.

胆固醇在决定生物膜的性质方面起着至关重要的作用。控制 细胞器中的胆固醇水平取决于尚不清楚的固醇转运机制。小说 已经开发了定量研究细胞器之间甾醇转运的方法，这导致了 细胞器间运输的精确模型。初步数据显示，一种广泛表达的甾醇- 受调节的可溶性甾醇转运蛋白STARD 4，在胆固醇的非囊泡转运中起重要作用， 并研究其作用和转运机制。STARD 4是一种复合物的重要组成部分， 固醇调节网络。最近的研究表明，STARD 4负责约25%的固醇转运 在U2 OS细胞中，两个主要池-质膜和内吞再循环室-之间。 STARD 4的作用也将在巨噬细胞中进行评估，巨噬细胞是一种胆固醇代谢起作用的细胞类型。 在动脉粥样硬化发展中的主要作用，以及在肝细胞模型HepG 2细胞中。初步 研究发现，STARD 4与磷脂酰肌醇磷酸（PIP）具有独特的相互作用。 当膜含有以下物质时，STARD 4从膜中提取甾醇的速度加快了近10倍 PI（4，5）P2，但PI（4，5）P2在接受来自STARD 4的固醇的膜中没有作用。相反地， 受体膜中的PI（3，5）P2或PI 5 P使转移加速10倍，当这些PIP与受体膜中的PI（3，5）P2或PI 5 P结合时， 供体膜。PI 3 P具有类似但较小的效果。STARD 4在晚期固醇转运中的作用 内体、脂滴和自噬体（即，具有良好受体PIP的细胞器）进行分析。 STARD 4与膜相互作用的分子动力学模拟正在确定潜在的 与PIP互动。这些位点突变的影响将在脂质体对脂质体固醇中进行测定 转运分析具有改变的PIP敏感性的突变STARD 4分子将在细胞中表达，以确定 对固醇运输和分布的影响。在初步研究中，通过分子动力学鉴定的突变体 通过脂质体测定中的测试证实，并且发现它们确实影响甾醇在 细胞器X射线晶体学已经确定了有和没有结合的人STARD 4之间的差异。 甾醇。NMR光谱学被用于分析STARD 4蛋白质动力学，并确定 与含有各种PIP的脂质相互作用。

项目成果

Membrane order in the plasma membrane and endocytic recycling compartment.

• DOI: 10.1371/journal.pone.0188041
• 发表时间: 2017
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Iaea DB;Maxfield FR
• 通讯作者: Maxfield FR

Role of STARD4 in sterol transport between the endocytic recycling compartment and the plasma membrane.

• DOI: 10.1091/mbc.e16-07-0499
• 发表时间: 2017-04-15
• 期刊: Molecular biology of the cell
• 影响因子: 3.3
• 作者: Iaea DB;Mao S;Lund FW;Maxfield FR
• 通讯作者: Maxfield FR

Phosphatidylinositol phosphates modulate interactions between the StarD4 sterol trafficking protein and lipid membranes.

• DOI: 10.1016/j.jbc.2022.102058
• 发表时间: 2022-07
• 期刊: JOURNAL OF BIOLOGICAL CHEMISTRY
• 影响因子: 4.8
• 作者: Zhang, Xiaoxue;Xie, Hengyi;Iaea, David;Khelashvili, George;Weinstein, Harel;Max, Frederick R.
• 通讯作者: Max, Frederick R.