Membrane Coupling and Dynamic Reorganization of Gag in Viral Budding

病毒出芽过程中膜耦合和堵嘴的动态重组

• 批准号: 8265158
• 负责人: Mathias Loesche
• 金额: $ 28.57万
• 依托单位: CARNEGIE-MELLON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-24 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8265158
• 关键词: 1-Phosphatidylinositol 3-Kinase; Address; Architecture; Binding; Biochemical; Capsid; Cell membrane; Cells; Color; Competitive Binding; Complement; Complex; Coupling; Cytoplasm; Daughter; Development; Disease; Electrostatics; Event; Face; Gagging; Genome; Genomics; Goals; Growth; HIV; HIV-1; Hydrophobic Interactions; Infection; Lateral; Lead; Length; Light; Lipid Bilayers; Lipids; Manuscripts; Measurement; Mediating; Membrane; Membrane Lipids; Membrane Proteins; Molecular; Molecular Conformation; Morphology; Murine leukemia virus; National Cancer Institute; Nature; Neutrons; Nucleic Acid Binding; Nucleic Acids; Nucleocapsid; Oncornaviruses; Oranges; Parents; Pathway interactions; Peptides; Phosphatidylinositol 4,5-Diphosphate; Play; Preparation; Process; Property; Protein Dynamics; Proteins; Qualifying; RNA; Recruitment Activity; Regulation; Reproduction; Research Personnel; Research Project Grants; Role; Series; Solid; Solutions; Sorting - Cell Movement; Specificity; Spectrum Analysis; Structure; Surface; Techniques; Testing; Therapeutic Agents; Therapeutic Intervention; Universities; Viral; Virion; Virus; Virus Replication; Work; computer studies; computerized tools; flexibility; gag Gene Products; molecular scale; novel therapeutics; particle; pathogen; prevent; protein expression; public health relevance; research study; single molecule; tool; viral RNA

DESCRIPTION (provided by applicant): The retroviral polyprotein Gag is the essential factor in a large number of viral pathogens, such as human immunodeficiency virus type 1 (HIV-1) and oncoviruses, that promotes the budding of progeny viral shells after binding to the host plasma membrane, leading to the formation of immature daughter particles which mature further into infectious viruses. A multitude of molecular interactions involved in membrane budding result in a complexity of the ensuing molecular reorganizations that impedes our understanding of these processes. This lack in understanding in turn prevents the targeting of this important step in the reproduction of virus particles with therapeutic interventions. Here, we focus on two hypotheses. (I) Gag binding to the lipid bilayer results from a hierarchical sequence of molecular interactions between the protein and the plasma membrane: Electrostatic steering of the membrane-binding domain, MA, to the bilayer surface; lipid-specific interaction of MA with the phosphatidylinositoldiphosphate PI(4,5)P2; and hydrophobic membrane insertion of a myristoyl anchor on MA's N-terminus. (II) In the full-length Gag polyprotein of HIV-1, the flexibility of the linker regions between distinct Gag domains is key for controlling the molecular reorganization that leads to membrane budding. We assembled a cross-disciplinary team of investigators who combine cutting-edge biophysical characterization capabilities of the structural and dynamic properties of proteins at membrane interfaces, located in Dr. Lashes' group at Carnegie Mellon University (CMU), with expertise in Gag protein expression, manipulation and characterization represented by Dr. Rein's group at the National Cancer Institute (NCI) which also has advanced capabilities in characterizing subcellular localization of protein constructs and the ultra- structural morphology of assembly products. These groups will work closely together to determine, in Aim 1, the factors that control the association of MA with membranes in binding studies, structural characterization with neutron scattering and quantitative measurements of the dynamics of protein association with lipid bilayers with correlation spectroscopy. With these tools, we also aim at understanding the mechanisms that lead to the recruitment of specific lipids into the viral shell. Under Aim 2, we characterize the origin and the implications of the conformational reorganization of full-length Gag at membrane surfaces in experimental and computational studies. Our team of investigators with its complementing expertise is uniquely qualified to boost our understanding of the molecular processes involved in viral shell formation. The public health relevance of this work lies in the development of a solid understanding of viral envelope formation in the replication step of HIV-1 virus and other retroviral pathogens. Its broader impact is to provide new techniques for the structural characterization of membrane proteins associated with lipid bilayers in their physiologically relevant, thermally disordered state. PUBLIC HEALTH RELEVANCE: The multiplication of viruses such as HIV in infected cells depends on the association of a viral protein, Gag, with the cell's plasma membrane which is consumed to form new viral shells in a complex sequence of molecular reorganizations. In this research project, we will determine the overall process that leads to the formation of viral shells because a thorough understanding of this process will be valuable for the development of new therapeutic strategies for interfering with virus formation in infected cells.

描述（由申请人提供）：逆转录病毒多蛋白Gag是大量病毒病原体（如人类免疫缺陷病毒1型（HIV-1）和癌病毒）的必需因子，它在与宿主质膜结合后促进子代病毒外壳出芽，形成未成熟的子颗粒，这些子颗粒进一步成熟为感染性病毒。参与膜出芽的众多分子相互作用导致随后的分子重组的复杂性，阻碍了我们对这些过程的理解。这种认识的缺乏反过来又阻碍了治疗干预措施针对病毒颗粒繁殖的这一重要步骤。在这里，我们关注两个假设。(1) Gag与脂质双分子层的结合是由分子相互作用的等级序列引起的