Control of Retrovirus Assembly by Membrane Lipid Thermodynamic Activity.

通过膜脂质热力学活性控制逆转录病毒组装。

• 批准号: 8865646
• 负责人: GERALD William FEIGENSON
• 金额: $ 29.34万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-15 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8865646
• 关键词: Acquired Immunodeficiency Syndrome; Address; Affect; Affinity; Behavior; Binding; Cell membrane; Cells; Charge; Cholesterol; Coupled; Disease; Drug Combinations; Electrostatics; Fluorescence Microscopy; Gagging; HIV; HIV-1; Health; Heterogeneity; Human; In Vitro; Invaded; Life; Life Cycle Stages; Lipid Binding; Lipids; Liquid substance; Measurement; Measures; Membrane; Membrane Lipids; Modeling; Morphology; Phase; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylserines; Phospholipids; Play; Process; Protein Binding; Proteins; RNA; Retroviridae; Role; Rous sarcoma virus; Site; Staging; Structural Protein; Surface; System; Thermodynamics; Viral; Viral Load result; Viral Proteins; Virion; Virus; Virus Assembly; Work; base; gag Gene Products; in vitro Model; membrane activity; membrane model; nanoscale

DESCRIPTION (provided by applicant): Viruses invade all forms of life, causing diseases in humans including HIV-AIDS. Although the variety of viruses is daunting, all enveloped viruses including those studied here, HIV-1 and Rous Sarcoma Virus (RSV), must associate with the cytoplasmic leaflet of plasma membranes. One difficulty for in vitro studies is the lack of good models of the plasma membrane cytoplasmic leaflet. Another difficulty is that real cells can have membrane heterogeneities on the tens of nanometer size scale on the opposed, exoplasmic leaflet. Both of these experimental issues are addressed in these proposed studies, with a multicomponent model cytoplasmic mixture that can be coupled to a phase-separated lipid mixture in an asymmetric bilayer. This project will explore how three aspects of membrane lipid mixing behavior are related to viral Gag protein binding and assembly: (1) How is the thermodynamic activity of membrane-bound phosphatidylserine controlled by lipid composition, and how is this PS activity connected to Gag binding? (2) How is the thermodynamic activity of Gag's other binding partner, PI(4,5)P2 controlled by the other membrane lipids, and in particular, what factors control the formation of PI(4,5)P2 domains? Are these domains the sites of Gag assembly? (3) The plasma membrane is asymmetric. How does the presence of a phase-separated leaflet that is coupled to the cytoplasmic leaflet change Gag binding and assembly? A theme of this work is that the tendency of membrane lipids to bind or react is described by their thermodynamic activity, and this activity is controlled by all the components of the mixture. This approach provides predictive power to describe the associations of viral Gag structural proteins with their lipid binding partners: Which membrane factors exert control over the interactions among membrane-bound viral Gag proteins? The overall strategy is to combine measurements of lipid thermodynamic activity with measurement of virus protein binding and assembly. Fluorescence microscopy is used to visualize domains of PI(4,5)P2, and to correlate these domains with measured Gag binding and Gag-Gag assembly into its viral lattice.

描述(申请人提供)：病毒侵入所有形式的生命，导致人类疾病，包括艾滋病毒-艾滋病。尽管病毒的种类令人望而生畏，但所有被包膜的病毒，包括这里研究的HIV-1和劳斯肉瘤病毒(RSV)，都必须与质膜的细胞质小叶联系在一起。体外研究的一个困难是缺乏良好的质膜细胞质小叶模型。另一个困难是，真正的细胞在相对的外质叶上可能具有数十纳米尺度的膜异质性。在这些拟议的研究中，这两个实验问题都得到了解决，多组分模型细胞质混合物可以耦合到不对称双层中的相分离的脂类混合物。本项目将探索膜脂混合行为与病毒Gag蛋白结合和组装的三个方面的关系：(1)膜结合磷脂酰丝氨酸的热力学活性如何受脂组成控制，这种PS活性如何与Gag结合有关？(2)Gag的另一个结合伙伴PI(4，5)P2的热力学活性如何受其他膜脂控制，特别是什么因素控制PI(4，5)P2结构域的形成？这些结构域是GAG的组装部位吗？(3)质膜不对称。连接到细胞质小叶上的相分离小叶的存在如何改变GAG的结合和组装？这项工作的一个主题是，膜脂结合或反应的趋势是由它们的热力学活性来描述的，这种活性受 混合物。这种方法提供了预测能力来描述病毒Gag结构蛋白与它们的脂结合伙伴之间的关联：哪些膜因素对膜结合的病毒Gag蛋白之间的相互作用起到控制作用？总体策略是将脂热力学活性的测量与病毒蛋白结合和组装的测量结合起来。荧光显微镜被用来显示PI(4，5)P2的结构域，并将这些结构域与测量到的Gag结合和Gag-Gag组装到其病毒晶格中相关联。