HIV-1 Gag Precursor Protein Interactions

HIV-1 Gag 前体蛋白相互作用

• 批准号: 10079388
• 负责人: ERIC W BARKLIS
• 金额: $ 50.11万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10079388
• 关键词: AIDS diagnosis; Acquired Immunodeficiency Syndrome; Binding; Binding Sites; Biochemical; C-terminal; Cell membrane; Ceramides; Cholesterol; Cytoplasmic Tail; Defect; Detergents; Development; Epidemic; Fostering; Foundations; Funding; Glycoproteins; Goals; HIV; HIV-1; Impairment; Intracellular Membranes; Investigation; Ligands; Lipids; Membrane; Membrane Microdomains; Methods; Modeling; Molecular Chaperones; Molecular Conformation; Monitor; Mutation; N-terminal; Phosphatidylinositol 4,5-Diphosphate; Phospholipids; Protein Precursors; Proteins; RNA; RNA Binding; Role; Signal Transduction; Site; Sphingomyelins; Structure; Transmembrane Domain; Variant; Viral; Virion; Virus; Virus Assembly; base; cell type; env Gene Products; gag Gene Products; insight; mutant; nanobodies; novel; novel strategies; therapy design; trafficking

Despite great advances in AIDS diagnosis and treatment, the continuing AIDS epidemic demands continuing efforts to understand all aspects of HIV replication and to develop new methods for its inhibition. In pursuit of these goals, we have sought to define the activities and interactions of the HIV-1 structural (Gag) proteins, with a specific focus on the N-terminal matrix (MA) domain. The Gag proteins initially are synthesized as precursor Gag (PrGag) proteins that are myristoylated at the N-terminus of MA, and MA domains target PrGag delivery to plasma membrane (PM) virus assembly sites virtue of preferential binding to the signaling phospholipid phosphatidylinositol-4,5-bisphosphate (PI[4,5]P2). Evidence also indicates that MA-RNA binding helps chaperone PrGag proteins to assembly sites, and that virus membranes are enriched for lipid raft constituents such as cholesterol, sphingomyelin, and ceramide. In addition to its trafficking role, MA has been shown to influence the incorporation of wild type (WT) HIV-1 envelope (Env) glycoprotein trimers into virus particles. Previous investigations have shown that HIV-1 Env proteins that carry cytoplasmic tail deletions (CT) in their transmembrane (TM; gp41) domains can be incorporated into virions in a fashion that is cell type-dependent, but MA-independent. In contrast, MA mutations that impair WT Env incorporation into virions have been identified. Moreover, other MA mutations have been shown to suppress Env incorporation defects imposed either by MA mutations, or Env CT mutations. Such observations imply that there could be direct MA-CT interactions, but proof has been lacking. During the past funding period, we have made significant progress in understanding how MA and Env proteins interact. We have shown that MA directly binds to Env CTs, and that binding depends on MA trimerization. We have demonstrated that C-terminal amphipathic helices of HIV-1 Env CTs are involved in MA binding, and that MA-CT binding is blocked by MA-RNA binding. We have discovered lipid composition changes that perturb WT Env activity, and have obtained novel evidence of CT processing. Using this as a foundation, we propose the characterization of MA-CT interactions and the roles of these interactions in HIV-1 replication. In particular, we will define how Env proteins associate with MA trimers and lattices so as to determine how WT Env proteins become incorporated into virus particles; and we will examine how Gag and Env proteins collaborate in virus particles to perform their functions. Our results will help clarify how HIV Gag and Env proteins cooperate, and will foster the development of novel approaches to interfere with HIV-1 replication.

尽管艾滋病的诊断和治疗取得了很大进展，但艾滋病的持续流行要求 继续努力了解艾滋病毒复制的方方面面，并制定抑制艾滋病毒复制的新方法。 为了实现这些目标，我们试图确定艾滋病毒-1结构的活动和相互作用 (GAG)蛋白，特别关注N-末端基质(MA)结构域。GAG蛋白最初是 合成前体GAG(PrGag)蛋白，在MA和MA的N端肉豆蔻酰化 靶向PrGag的结构域能优先结合到质膜(PM)病毒组装部位 信号转导通路磷脂酰肌醇-4，5-二磷酸(PI[4，5]P2)。证据还表明， MA-RNA结合帮助伴侣PrGag蛋白到组装位置，并且病毒膜是 富含脂筏成分，如胆固醇、神经鞘蛋白和神经酰胺。除了它的 运输作用，MA已被证明影响野生型(WT)HIV-1外膜(Env)的加入 糖蛋白三聚体形成病毒颗粒。先前的研究表明，HIV-1包膜蛋白 携带跨膜(TM；gp41)结构域中的细胞质尾部缺失(CT)可被整合到 病毒粒子依赖于细胞类型，但不依赖于MA。相比之下，MA突变会损害 WT Env掺入病毒粒子已被鉴定。此外，其他MA突变已被证明是 抑制由MA突变或Env CT突变造成的Env掺入缺陷。是这样的 观察表明，可能存在MA-CT的直接相互作用，但一直缺乏证据。在.期间 在过去的资助期间，我们在了解MA和Env蛋白如何相互作用方面取得了重大进展。 我们已经证明了MA直接与环境CTs结合，这种结合依赖于MA的三聚化。我们有 证明HIV-1 Env CTs的C末端两亲性螺旋参与MA结合，并且 MA-CT结合被MA-RNA结合阻断。我们已经发现脂质成分的变化会扰乱 WT Env活性，并获得了CT加工的新证据。以此为基础，我们 提出MA-CT相互作用的特征以及这些相互作用在HIV-1复制中的作用。 特别是，我们将定义Env蛋白如何与MA三聚体和晶格相关联，从而确定如何 WT Env蛋白被整合到病毒颗粒中；我们将研究Gag和Env蛋白是如何 在病毒颗粒中进行协作以执行它们的功能。我们的结果将有助于阐明HIV Gag和Env是如何 蛋白质相互协作，并将促进干扰HIV-1复制的新方法的开发。