GENETIC ANALYSIS OF HERPES VIRUS NEUROTROPISM AND ENCEPHALITIS

疱疹病毒向神经性和脑炎的遗传分析

• 批准号: 8169768
• 负责人: JENNIFER Hart LAVAIL
• 金额: $ 0.18万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-12 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8169768
• 关键词: Affinity; Antibodies; Axonal Transport; Biochemical; Capsid; Capsid Proteins; Cell model; Computer Retrieval of Information on Scientific Projects Database; Coupled; Cytoplasm; DNA; Encephalitis; Funding; Gel; Grant; Herpesviridae; Herpesvirus 1; Immunohistochemistry; Institution; Label; Mass Spectrum Analysis; Modeling; Motor Neurons; Mus; Neurons; Neurotropism; Nucleocapsid; Presynaptic Terminals; Process; Proteins; Research; Research Personnel; Resources; Retinal Ganglion Cells; Sepharose; Silver Staining; Source; United States National Institutes of Health; Viral; Viral Envelope Proteins; Viral Proteins; Virus; Visual system structure; anterograde transport; genetic analysis; mutant; neuronal cell body; therapeutic target; tool; transmission process; vesicle-associated membrane protein; viral DNA

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. A critical step in the transmission of Herpes simplex virus type 1 virus (HSV) from one infected neuron to the next is the polarized anterograde axonal transport of viral DNA from the host infected nerve cell body to the axon terminal for subsequent release. How the virus is transported and what viral proteins are necessary are long-standing questions. Using an HSV mutant virus that lacks expression of the Us9 protein and the infected murine retinal ganglion cell model, we found that HSV Us9 protein is necessary specifically for long distance anterograde axonal transport of viral capsid and DNA. It is unnecessary for anterograde transport of viral envelope proteins or for retrograde axonal transport of HSV. Using an immunoaffinity matrix of Us9 antibody coupled to Sepharose beads, we co-concentrated Us9 and VP5, the major capsid protein. This biochemical evidence suggests a mechanism by which the capsid transport depends on an association with Us9 protein. This association was further confirmed with EM immunohistochemistry in which Us9 antibody labeled unenveloped capsids in wild-type virus infected retinal ganglion cell cytoplasm. We conclude that efficient axonal transport of HSV DNA and capsid depends on expression of Us9 protein and does not require the traditional membrane vesicle proteins that are associated with many host cell motors. We shall affinity purify proteins that associate with Us9 on the affinity matrix and then separate them on PAGE. The proteins will be silver stained and cut out of the gels for identification using mass spectroscopy for identification. As a model of non-vesicular transport, the anterograde axonal transport of HSV nucleocapsid offers a new biochemical tool for understanding this functional process in normal neurons. Furthermore, Us9 protein is a potential therapeutic target against the spread of HSV in the visual system.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 从一个感染的神经元到下一个的单纯性1型病毒（HSV）传播的关键步骤是，病毒DNA从宿主感染的神经细胞体到轴突末端的偏光顺行轴突转运，以便随后释放。 该病毒的运输方式以及需要哪种病毒蛋白是长期存在的问题。 使用缺乏US9蛋白表达和感染的鼠视网膜神经节细胞模型的HSV突变病毒，我们发现HSV US9蛋白对于病毒capsid和DNA的长距离顺行轴突运输是必需的。 对于病毒包膜蛋白的顺行转运或HSV的逆行轴突转运是不必要的。 使用与Sepharose珠相连的US9抗体的免疫亲和力基质，我们将US9和VP5（主要的capsid蛋白）共含有。 该生化证据表明了一种机制，可以使衣壳转运取决于与US9蛋白的关联。 通过EM免疫组织化学进一步证实了这种关联，其中US9抗体在野生型病毒感染了视网膜神经节细胞细胞质中标记了未开发的衣壳。 我们得出的结论是，HSV DNA和CAPSID的有效轴突转运取决于US9蛋白的表达，并且不需要与许多宿主细胞电机相关的传统膜囊泡蛋白。 我们应将与亲和力矩阵上US9关联的蛋白质相关，然后在页面上将它们分开。 蛋白质将是银色染色并从凝胶中切出的，以使用质谱识别识别蛋白质。 作为非挥发性转运的模型，HSV核素的顺行轴突转运提供了一种新的生化工具，用于理解正常神经元中的这种功能过程。 此外，US9蛋白是针对HSV在视觉系统中扩散的潜在治疗靶标。