HIV-1 Gag Precursor Protein Interactions

HIV-1 Gag 前体蛋白相互作用

• 批准号: 10176400
• 负责人: ERIC W BARKLIS
• 金额: $ 49.02万
• 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/10176400
• 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.

尽管艾滋病诊断和治疗取得了巨大进步，但持续的艾滋病流行仍需要 继续努力了解艾滋病毒复制的各个方面并开发新的抑制方法。 为了实现这些目标，我们试图定义 HIV-1 结构的活动和相互作用。 (Gag) 蛋白，特别关注 N 端基质 (MA) 结构域。 Gag 蛋白最初是 合成为前体 Gag (PrGag) 蛋白，在 MA 和 MA 的 N 末端被肉豆蔻酰化 结构域通过优先结合将 PrGag 递送至质膜 (PM) 病毒组装位点 信号磷脂磷脂酰肌醇-4,5-二磷酸 (PI[4,5]P2)。证据还表明 MA-RNA 结合有助于伴侣 PrGag 蛋白到达组装位点，并且病毒膜 富含胆固醇、鞘磷脂和神经酰胺等脂筏成分。除了它的 MA 已被证明可以影响野生型 (WT) HIV-1 包膜 (Env) 的掺入 糖蛋白三聚体转化为病毒颗粒。先前的研究表明，HIV-1 Env 蛋白 在其跨膜（TM；gp41）结构域中携带细胞质尾部缺失（CT），可以将其整合到 病毒粒子以细胞类型依赖性但与 MA 无关的方式产生。相反，MA突变会损害 已鉴定出 WT Env 掺入病毒颗粒中。此外，其他 MA 突变已被证明 抑制由 MA 突变或 Env CT 突变造成的 Env 掺入缺陷。这样的 观察结果表明 MA-CT 可能存在直接相互作用，但缺乏证据。期间 在过去的资助期间，我们在理解 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 MA 直接与 Env CT 结合，并且这种结合取决于 MA 三聚化。我们有 证明 HIV-1 Env CT 的 C 端两亲性螺旋参与 MA 结合，并且 MA-CT 结合被 MA-RNA 结合阻断。我们发现脂质成分的变化会扰乱 WT Env 活性，并获得了 CT 处理的新证据。以此为基础，我们 提出了 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 三聚体和晶格结合，以确定如何 WT Env 蛋白被整合到病毒颗粒中；我们将研究 Gag 和 Env 蛋白如何 病毒颗粒相互协作以发挥其功能。我们的结果将有助于阐明 HIV Gag 和 Env 如何 蛋白质相互配合，将促进干扰 HIV-1 复制的新方法的开发。