Mechanism of APOBEC3G-Mediated Hypermutation and Inhibit

APOBEC3G 介导的超突变和抑制机制

• 批准号: 7064431
• 负责人: VINAY K. PATHAK
• 金额: --
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7064431
• 关键词: Retroviridae; aminoacid; aminohydrolases; cell line; cytidine; deoxycytidine; enzyme mechanism; gene expression; gene mutation; host organism interaction; human immunodeficiency virus 1; intermolecular interaction; protein binding; protein degradation; virion; virus genetics; virus protein; virus replication

HIV-1 and other retroviruses occasionally undergo a high rate of G-to-A substitutions, a phenomenon named hypermutation. HIV-1 genomes that fail to express the accessory protein Vif cannot replicate in primary cells or "nonpermissive" cell lines but can replicate in "permissive" cell lines. APOBEC3G, the dominant-acting host restriction factor responsible for the nonpermissive phenotype, is a cytidine deaminase that is packaged into HIV-1 virions in the absence of Vif and deaminates deoxycytidines in minus-strand DNA to deoxyuridines, resulting in massive G-to-A hypermutation and abrogation of viral replication. HIV-1 Vif binds to APOBEC3G and induces its proteosomal degradation in the virus producer cells, suppressing its virion incorporation and restoring viral replication. Human APOBEC3G is degraded in the presence of Vif but simian APOBEC3G is resistant to Vif; using mutational analysis, we and others recently showed that a single amino acid (D128) is responsible for the species specificity of APOBEC3G proteins. The D128 residue either directly interacts with Vif or is involved in conformational changes that occur upon Vif binding. To gain insights into the mechanism by which APOBEC3G inhibits HIV-1 replication, we recently developed a sensitive cytidine deamination assay using scintillation proximity beads. Using this assay, we demonstrated that interactions with viral and nonviral RNAs that are packaged are sufficient for APOBEC3G virion incorporation and that interactions with viral proteins are not essential for virion incorporation. Our future goals are to elucidate the structure and function of APOBEC3G, identify other host proteins that are critical for APOBEC3G-mediated inhibition of HIV-1 replication, define the nature of the Vif-APOBEC3G interactions, and develop agents that interfere with Vif-APOBEC3G interactions as potential antiviral agents.

HIV-1和其他逆转录病毒偶尔会经历高率的G-TO替换率，这一现象称为HyperMunt。无法表达辅助蛋白VIF的HIV-1基因组无法在原代细胞或“非允许”细胞系中复制，但可以在“允许”细胞系中复制。 APOBEC3G是负责非耐药表型的主要作用宿主限制因子，是一种细胞丁胺脱氨酶，在没有VIF的情况下将其包装到HIV-1病毒体中，并脱氧于脱氧尿素中的脱氧于氧基尿素中的脱氧胞苷，导致甲氧基尿尿素的脱氧尿素，从而导致量大的高度逐渐渗透性，导致高度逐渐变色的含量。 HIV-1 VIF与APOBEC3G结合，并在病毒生产细胞中诱导其蛋白质体降解，从而抑制其病毒颗粒掺入并恢复病毒复制。在VIF的存在下，人Apobec3g降解，但Simian apobec3g对VIF具有抵抗力。使用突变分析，我们和其他人最近表明，单个氨基酸（D128）负责APOBEC3G蛋白的物种特异性。 D128残基要么直接与VIF相互作用，要么参与VIF结合时发生的构象变化。为了洞悉APOBEC3G抑制HIV-1复制的机制，我们最近使用闪烁接近珠开发了一种敏感的胞苷脱氨基测定法。使用该测定法，我们证明了包装的病毒和非病毒RNA的相互作用足以用于Apobec3g病毒率掺入，并且与病毒蛋白的相互作用对于Virion掺入并不是必需的。我们的未来目标是阐明Apobec3g的结构和功能，识别其他宿主蛋白，这些宿主蛋白对于APOBEC3G介导的HIV-1复制抑制作用至关重要，定义了VIF-APOBEC3G相互作用的性质，并发展与VIF-APOBEC3G相互作用相互作用作为潜在抗病毒剂的抗病毒剂。