GENETICS OF PRIMATE 'D' TYPE RETROVIRUSES

灵长类“D”型逆转录病毒的遗传学

• 批准号: 8357425
• 负责人: Eric Hunter
• 金额: $ 7.43万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8357425
• 关键词: AIDS/HIV problem; Actins; Biological Assay; Capsid; Cell membrane; Cells; Complex; Cytoskeleton; Funding; Gagging; Genetic; Glycoproteins; Grant; Hour; Life; Microtubules; Minor; Motor; Movement; National Center for Research Resources; Nocodazole; Pathway interactions; Pericentriolar Regions; Physiologic pulse; Primates; Principal Investigator; Production; Recovery; Research; Research Infrastructure; Resources; Role; Site; Source; Tubular formation; Type D Retrovirus; United States National Institutes of Health; Viral; Virion; Virus; anterograde transport; cellular imaging; cost; dynein light chain; retrograde transport; trafficking

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Objectives: define (1) M-PMV Gag interactions with the cytoskeletal motor machinery, (2) role of viral and cellular components in transporting capsids from their assembly site to the plasma membrane, (3) capsid interactions with the cellular ESCRT machinery, and (4) the myristyl switch mechanism involved in budding. Efficient transport of pre-assembled capsids to the cell membrane requires a functional vesicular trafficking pathway and envelope (Env) glycoprotein. Transport may occur along a tubular-vesicular pathway induced by Env or another viral component, or these interactions may initiate Gag transport along other trafficking networks. We show Gag interacts in a CTRS-dependent manner with cellular Tctex-1, a light chain of the dynein motor complex, suggesting a microtubule-dependent retrograde transport to the pericentriolar region for capsid assembly. Microtubules might also explain anterograde transport of pre-assembled capsids to the plasma membrane. In M-PMV-infected CMMT cells, pulse-chase assays revealed a nocodazole induced 1-hour delay in virus production compared to untreated controls. Actin disruption led to a minor delay in virion release, while IF disruption had negligible effects. Live cell imaging showed that following microtubule disruption, an initial reduction in linear movement of capsids containing GFP-tagged Gag occurred, followed by a recovery of velocities. Nocodazole treatment led to a cessation in Env transport and deficient Env incorporation in released virions. Importantly, a continued albeit delayed release of virions was observed from cells lacking all three functional cytoskeleton components, suggesting a cytoskeleton-independent mode of transport. Studies are underway to alternate transport pathways. These studies could have implications for HIV/AIDS research.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的主要研究者可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 目标：定义 (1) M-PMV Gag 与细胞骨架运动机制的相互作用，(2) 病毒和细胞成分在将衣壳从其组装位点运输到质膜中的作用，(3) 衣壳与细胞 ESCRT 机制的相互作用，以及(4)参与出芽的肉豆蔻基开关机制。将预组装衣壳有效转运至细胞膜需要功能性囊泡运输途径和包膜 (Env) 糖蛋白。运输可能沿着 Env 或其他病毒成分诱导的管状囊泡途径发生，或者这些相互作用可能启动沿着其他运输网络的 Gag 运输。我们发现 Gag 以 CTRS 依赖性方式与细胞 Tctex-1（动力蛋白运动复合体的一条轻链）相互作用，表明微管依赖性逆行运输到中心粒周围区域进行衣壳组装。微管还可以解释预组装衣壳向质膜的顺行运输。在 M-PMV 感染的 CMMT 细胞中，脉冲追踪分析显示，与未处理的对照相比，诺考达唑诱导病毒产生延迟 1 小时。肌动蛋白破坏导致病毒颗粒释放略有延迟，而 IF 破坏的影响可以忽略不计。活细胞成像显示，微管破坏后，含有 GFP 标记 Gag 的衣壳的线性运动最初减少，随后速度恢复。诺考达唑治疗导致 Env 运输停止，释放的病毒颗粒中 Env 掺入不足。重要的是，在缺乏所有三种功能性细胞骨架成分的细胞中观察到病毒粒子持续但延迟的释放，这表明存在一种独立于细胞骨架的运输模式。正在研究替代运输途径。这些研究可能对艾滋病毒/艾滋病研究产生影响。