Allostery and Hijacking of Host Membrane Traffic by HIV-1 Accessory Proteins

HIV-1 辅助蛋白对宿主膜运输的变构和劫持

• 批准号: 10669213
• 负责人: James H Hurley
• 金额: $ 64.68万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-18 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10669213
• 关键词: Acquired Immunodeficiency Syndrome; Adaptor Protein Complex 2; Adaptor Signaling Protein; Affect; Alleles; Antiviral Agents; Automobile Driving; Basic Science; Binding Sites; Biochemistry; Biophysics; CD28 gene; CD3 Antigens; CD8B1 gene; CXCR4 gene; Cell Surface Proteins; Cell membrane; Cell surface; Cells; Clathrin; Clathrin-Coated Vesicles; Complement; Complex; Cryoelectron Microscopy; Cytosol; Detergents; Down-Regulation; Family; Funding; Goals; HIV; HIV-1; Host Defense; Human; Image; Immune system; Immunity; Infection; Integral Membrane Protein; Lipids; Location; Lysosomes; Mediating; Membrane; Membrane Lipids; Membrane Protein Traffic; Monomeric GTP-Binding Proteins; Mutation; Pathway interactions; Pattern; Phenotype; Process; Production; Proteins; Reporting; Research Proposals; Role; Route; SIV; Seminal; Signal Transduction; Site; Sorting; Structure; Substrate Interaction; Tail; Viral; Viral Antigens; Virion; Virus; design; follow-up; insight; latent virus activation; member; prevent; protein complex; reactivation from latency; receptor; reconstitution; trans-Golgi Network; viral fitness

PROJECT SUMMARY Nef is an HIV-1 accessory protein whose function is to undermine host defenses. Long term infection with HIV strains bearing defective nef alleles leads to AIDS only over many years, suggesting Nef could be targeted as part of functional cure strategies. This basic research proposal seeks will elucidate the mechanisms of action of Nef. Nef targets include CD3, CD4, CD8, CD28, CXCR4, MHC-I, SERINC3/5, and for HIV-1 group O and SIV, BST2/tetherin. Nef substrates are downmodulated by via the clathrin-coated vesicle (CCV) pathway. Nef does not interact directly with clathrin, but rather with various members of a family of heterotetrameric adaptors known as the adaptor protein (AP) complexes, AP-1 and AP-2. The ability of the human immune system to detect and kill virally infected cells relies on proper presentation of viral antigens on the cell surface by MHC-I complexes. Nef subverts this process by promoting MHC-I complex downregulation by hijacking AP-1 and its associated small GTPase Arf1 at the TGN. MHC-I contains an incomplete version of the normal Tyr-based sorting motif. Nef complements this defective motif and converts MHC-I into a substrate for AP-1 mediated sorting to the lysosome for degradation. Structures of Nef assembled with AP-1 and the MHC-I cytosolic tail in solution suggested that Nef promotes the assembly of hexagonal lattices whose symmetry matches that of clathrin. Now, the previous solution studies will be followed up by reconstitution and structure determination of Nef, MHC-I tail, AP-1 and Arf1 in their functional setting on lipid membranes. Downmodulation of CD28 by Nefs is conserved across SIV and HIV and is mediated by AP-2. CD28 downmodulation phenotypes of Nef mutations follow a distinct pattern from other receptors, and the structural basis for this mode of CD28 is unknown. The structure of the CD28:HIV-1 Nef:AP-2 complex will be determined and leveraged to design mutations that uniquely perturb the CD28 binding site. Of the many host substrates of Nef, the most significant for viral infectivity are the multipass integral membrane proteins SERINC3 and 5. The SERINC binding site appears, on the basis of Nef phenotypes, to overlap with the site used by SIVsmm Nef to downmodulate simian tetherin, but is otherwise distinct from the known locations of CD3, CD4, and MHC-I sites. SERINCs do not share any obvious motifs with other substrates. SERINCs have been purified in monodisperse form suitable for structure determination. The cryo-EM structure of lipid- or detergent embedded SERINC3 or 5 in complex with AP-2 and HIV-1 Nef will be determined, completing a major goal in the field.

项目摘要 Nef是一种HIV-1辅助蛋白，其功能是破坏宿主防御。长期感染HIV 携带有缺陷nef等位基因的菌株仅在多年后才导致艾滋病，这表明Nef可以作为靶点， 功能性治疗策略的一部分。这项基础研究计划旨在阐明 的Nef。Nef靶点包括CD 3、CD 4、CD 8、CD 28、CXCR 4、MHC-I、SERINC 3/5，并且对于HIV-1的O组和 SIV，BST 2/栓系蛋白。Nef底物通过网格蛋白包被囊泡（CCV）途径下调。NEF 不直接与网格蛋白相互作用，而是与异源四聚体衔接子家族的各种成员相互作用 称为衔接蛋白（AP）复合物，AP-1和AP-2。人类免疫系统的能力， 检测和杀死病毒感染的细胞依赖于通过MHC-I在细胞表面上正确呈递病毒抗原 配合物Nef通过劫持AP-1及其受体来促进MHC-I复合体下调，从而颠覆了这一过程。 在TGN的相关的小GTdR Arf 1。MHC-I包含正常Tyr基 排序基序Nef补充了这个缺陷的基序，并将MHC-I转化为AP-1介导的底物。 分选到溶酶体中降解。Nef与AP-1和MHC-I胞质尾组装的结构在人肝癌细胞中的表达 解决方案表明，Nef促进了六方晶格的组装，其对称性与 网格蛋白。现在，之前的溶液研究将通过以下步骤进行重构和结构测定： Nef、MHC-I尾、AP-1和Arf 1在脂质膜上的功能设置。Nefs下调CD 28 在SIV和HIV中是保守的，并由AP-2介导。Nef的CD 28下调表型 突变遵循与其他受体不同的模式，CD 28这种模式的结构基础是 未知将确定CD 28：HIV-1 Nef：AP-2复合物的结构，并利用其设计 独特地干扰CD 28结合位点的突变。在Nef的许多宿主底物中， 用于病毒感染性的是多通道整合膜蛋白SERINC 3和5。SERINC结合位点 根据Nef表型，似乎与SIVsmm Nef用于下调 猴系链蛋白，但在其他方面不同于已知的CD 3、CD 4和MHC-I位点的位置。SERINC确实 不与其他基材共享任何明显的基序。SERINC已被纯化为单分散形式 适用于结构测定。脂质或去污剂包埋的SERINC 3或5的冷冻-EM结构， 将确定与AP-2和HIV-1 Nef的复合物，完成该领域的主要目标。

项目成果

HIV-1 Nef hijacks clathrin coats by stabilizing AP-1:Arf1 polygons.

• DOI: 10.1126/science.aac5137
• 发表时间: 2015-10-23
• 期刊: Science (New York, N.Y.)
• 作者: Shen QT;Ren X;Zhang R;Lee IH;Hurley JH
• 通讯作者: Hurley JH

Clathrin-associated AP-1 controls termination of STING signalling.

• DOI: 10.1038/s41586-022-05354-0
• 发表时间: 2022-10
• 期刊: NATURE
• 影响因子: 64.8
• 作者: Liu, Ying;Xu, Pengbiao;Rivara, Sophie;Liu, Chong;Ricci, Jonathan;Ren, Xuefeng;Hurley, James H.;Ablasser, Andrea
• 通讯作者: Ablasser, Andrea