Clarifying the role of the cytoplasmic dynein motor complex in polyomavirus nuclear entry.

阐明细胞质动力蛋白运动复合物在多瘤病毒核进入中的作用。

• 批准号: 9758812
• 负责人: Chelsey Cierra Spriggs
• 金额: $ 6.16万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9758812
• 关键词: Adaptor Signaling Protein; Affinity; Antiviral Agents; Binding; Capsid; Cell Nucleus; Cell physiology; Cell surface; Cells; Complex; Cytosol; DNA Tumor Viruses; DNA Viruses; Disease; Dynein ATPase; Endoplasmic Reticulum; Event; Genetic; Genetic Transcription; Genome; Human; Immunocompromised Host; Impairment; Individual; Infection; Intracellular Transport; Lead; Life Cycle Stages; Lytic Phase; Mammalian Cell; Membrane; Merkel Cells; Microtubules; Motor; Motor Activity; Nuclear; Nuclear Pore Complex; Pathway interactions; Polyomavirus; Polyomavirus Infections; Prevention; Process; Reporting; Research Proposals; Role; Simian virus 40; Skin Carcinoma; Specificity; Structure; Testing; Viral; Viral Genome; Virion; Virus; Virus Replication; base; cell transformation; ds-DNA; dynactin; experimental study; human disease; insight; knock-down; mechanical force; metaplastic cell transformation; particle; protein complex; recruit; virus core

Project Summary/Abstract Polyomaviruses (PyVs) are small DNA tumor viruses that cause debilitating human disease, especially in immunocompromised individuals. To infect cells, these non-enveloped viruses must transport to the host nucleus where transcription and replication of the viral genome lead to lytic infection or cellular transformation. During entry, PyV sorts from the cell surface to the endoplasmic reticulum (ER) where it penetrates the ER membrane to reach the cytosol. From here, the virus is disassembled in order to cross the narrow nuclear pore complex (NPC) and enter the nucleus. PyV transport from the cytosol to the nucleus is an important, yet enigmatic, step in infection. In mammalian cells, intracellular transport to the nucleus is facilitated largely by the cytoplasmic motor dynein, which moves cargo along microtubules towards the center of the cell. Using the prototypic PyV, simian virus 40 (SV40), which shares both structural and genetic organization with human PyVs as well as the same infectious life cycle, we recently reported that dynein motor activity is required for viral disassembly and nuclear arrival of the virus. The exact mechanisms by which dynein promotes this process are unknown. Processive dynein activity requires a three-protein complex composed of the dynein motor, dynactin activator, and an adaptor protein, which confers cargo specificity. Preliminary experiments reveal that in addition to dynein and dynactin, the bicaudal D2 (BICD2) cargo adaptor is also important for SV40 infection. The knockdown of BICD2 significantly impairs SV40 disassembly in the cytosol as well as its nuclear arrival. Moreover, BICD2 interacts directly with the virus and promotes its release at the NPC. In addition to activation by cargo adaptors, dynein activity can also be regulated by LIS1 with either NUDE or NDEL, co-factors that serve to anchor the motor to its microtubule track. Interestingly, we found that LIS1 and NDEL are also essential for SV40 disassembly and infection. This research proposal aims to define the role of dynein motor complex activators and regulators in the PyV entry pathway (Aim1), and whether the virus is directly capable of regulating this cellular process to complete its life cycle (Aim2). We hypothesize that SV40 recruits the dynein-dynactin-BICD2 (DDB) complex in the cytosol, which in turn transports the viral particle into the nucleus. Because the intact virus is too large to transport across the NPC, we further postulate that the coordinated action of dynein activators (BICD2) and regulators (LIS1/NDEL) produces a mechanical force that is strategically harnessed by the virus to generate a smaller core virus that can enter NPC. Upon completion of these studies, our findings will illuminate key steps in the PyV entry pathway and identify potential anti-viral targets for the prevention and treatment of PyV infection and disease.

项目摘要/摘要 多瘤病毒（PYVS）是导致人类疾病的小型DNA肿瘤病毒，尤其是在 免疫功能低下的个体。要感染细胞，这些非发育的病毒必须运输到宿主 病毒基因组的转录和复制导致裂解感染或细胞转化的细胞核。 在进入期间，PYV从细胞表面到内质网（ER），在那里穿透ER 膜到达细胞质。从这里开始，该病毒被解散以越过狭窄的核孔 复合物（NPC）并进入核。 PYV从细胞质到核的转运是重要的，但 神秘，感染的一步。在哺乳动物细胞中，细胞内转运向细胞核的转运主要由 细胞质运动动力蛋白，该动力蛋白沿着微管移动到细胞中心。使用 原型PYV，Simian病毒40（SV40），它与人类共享结构和遗传组织 PYV和相同的感染生命周期，我们最近报告说，Dynein运动活动需要 病毒拆卸和病毒的核到来。动力蛋白促进这一点的确切机制 过程未知。进程动力蛋白活性需要由动力蛋白组成的三蛋白复合物 电动机，dynactin激活剂和衔接蛋白，赋予货物特异性。初步实验 揭示除了Dynein和Dynactin之外，Bicauaudal D2（BICD2）货物适配器对 SV40感染。 BICD2的敲低显着损害了细胞质中的SV40拆卸 核到来。此外，BICD2与病毒直接相互作用，并在NPC促进其释放。在 除货物适配器的激活外，Dynein活性也可以由Lis1调节，裸体或 NDEL，用于将电动机锚定在其微管轨道上的联合因素。有趣的是，我们发现Lis1和 NDEL对于SV40拆卸和感染也是必不可少的。该研究建议旨在定义 PYV进入途径中的Dynein Motor复合激活剂和调节剂（AIM1），以及该病毒是否为 直接能够调节这种细胞过程以完成其生命周期（AIM2）。我们假设SV40 在细胞质中募集动力蛋白二奈霉素-BICD2（DDB）复合物，进而将病毒颗粒传输到 核。因为完整的病毒太大而无法跨NPC运输，所以我们进一步假设 动力蛋白激活剂（BICD2）和调节剂（LIS1/NDEL）的协调作用产生一种机械力，该力量 病毒在战略上利用了可以输入NPC的较小核心病毒。完成后 这些研究，我们的发现将阐明PYV进入途径中的关键步骤，并确定潜在的抗病毒 预防和治疗PYV感染和疾病的靶标。