Imaging of HTLV-1 by cryo-CLEM

通过冷冻 CLEM 进行 HTLV-1 成像

• 批准号: 10868844
• 负责人: Louis M Mansky
• 金额: $ 8.42万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-16 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10868844
• 关键词: 2019-nCoV; Address; Adult T-Cell Leukemia/Lymphoma; Behavior; Biogenesis; Biomedical Engineering; Capsid; Cell Communication; Cell Culture Techniques; Cells; Cellular biology; Cryoelectron Microscopy; Development; Disease; Electron Microscope; Electron Microscopy; Engineering; Etiology; Event; Eye; Fluorescence Microscopy; Goals; Human; Human Characteristics; Human T-Cell Leukemia Viruses; Human T-lymphotropic virus 1; Image; Imaging technology; Individual; Infection; Inflammatory; Knowledge; Lasers; Malignant Neoplasms; Morphology; Nature; Oral; Organ; Pattern; Preparation; Proteins; Pulmonary Pathology; Reproducibility; Research; Research Project Grants; Resolution; Retroviridae; Role; Sampling; Skin; Spinal Cord Diseases; Syndrome; Technology; Testing; Thyroid Gland; Tissues; Tropical Spastic Paraparesis; Viral; Virion; Virus; Virus Assembly; Virus Diseases; Virus Replication; experimental study; fluorescence imaging; high resolution imaging; human imaging; imaging study; improved; insight; instrument; light microscopy; live cell imaging; microscopic imaging; mutant; new technology; novel; particle; permissiveness; quantitative imaging; success; superresolution microscopy; transmission process; viral transmission; virological synapse; virology

RESEARCH PROJECT Abstract Human T-cell leukemia virus (HTLV-1) has been estimated to infect 15-20 million individuals worldwide and is known to be the etiological agent of an adult T-cell leukemia/lymphoma (ATLL), an inflammatory disease syndrome known as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), and pathologies of the lung, skin, eyes, and thyroid gland. HTLV-1 is notorious for being extremely difficult to propagate in cell culture, which has prohibited rigorous analyses of virus replication, including the steps involved in retrovirus assembly. HTLV-1 spread is known to be heavily reliant on virus infection involving cell-to-cell contacts that form what is termed the virological synapse (VS), which represents the primary means for virus spread, including events associated with oral transmission. While HTLV-1 has been previously studied in regard to virus spread via cell-cell contacts, a significant knowledge gap exists regarding the nature of virus particle assembly and transmission via the VS. In general, virus particle spread through cell-cell contacts increases the likelihood of an infection event of virus particles that may possess low particle infectivity in cells without formation of VS. Previous studies have indicated that a low proportion of mature HTLV-1 particles possess an intact capsid core, suggesting that aberrant particle morphology could help to explain the poorly infectious nature of cell-free HTLV- 1. In order to address the current knowledge gap in the field, we propose in this exploratory application to develop workflows for state-of-the-art bioengineering and quantitative imaging technologies that hold high promise in being applied to the efficient study of HTLV-1 particle assembly and spread at the VS. First, we will develop a workflow for the use of cell micropatterning technology in order to reproducibly and efficiently create cell-cell contacts and investigate the role virus budding in virus spread. Second, we propose to establish an efficient workflow in which we can view cell-cell contacts utilizing high-resolution cryo-correlative light and electron microscopy in order to investigate the role of host cell proteins in virus assembly at cell-cell contacts. Development of these workflows will allow for quantitative analysis of virus particle biogenesis at cell-cell contacts. These technologies have broad applicability in virology and the success of this research will be applicable to a variety of questions regarding virus replication and virus-host cell interactions.

研究项目 摘要 据估计，人类T细胞白血病病毒（HTLV-1）感染了全世界1500万至2000万个体， 已知是成人T细胞白血病/淋巴瘤（ATLL）的病原体， 被称为HTLV-1相关脊髓病/热带痉挛性轻瘫（HAM/TSP）的综合征和病理 肺皮肤眼睛和甲状腺HTLV-1因在细胞中极难繁殖而臭名昭著 文化，这已经禁止严格分析病毒复制，包括逆转录病毒的步骤， 组装件.已知HTLV-1传播严重依赖于涉及细胞间接触的病毒感染， 病毒学突触（VS）是病毒传播的主要途径，包括 与口腔传播有关的事件。虽然HTLV-1以前已经研究过病毒传播 通过细胞-细胞接触，关于病毒颗粒组装的性质存在显著的知识缺口， 一般来说，通过细胞-细胞接触传播的病毒颗粒增加了病毒传播的可能性。 在细胞中可能具有低颗粒感染性而不形成VS的病毒颗粒的感染事件。 研究表明低比例的成熟HTLV-1颗粒具有完整的衣壳核心， 这表明异常的颗粒形态可能有助于解释无细胞HTLV的感染性差的性质， 1.为了解决目前在该领域的知识差距，我们建议在这个探索性的应用程序开发 最先进的生物工程和定量成像技术的工作流程， 应用于HTLV-1粒子在VS的组装和扩散的有效研究。首先，我们将开发一个 使用细胞微图案化技术以便可再现地和有效地创建细胞-细胞 接触和调查病毒出芽在病毒传播中的作用。第二，我们建议建立一个有效的 我们可以利用高分辨率的冷冻相关光和电子来观察细胞与细胞之间的接触 在显微镜下观察以研究宿主细胞蛋白在细胞-细胞接触的病毒组装中的作用。 这些工作流程的开发将允许在细胞-细胞上定量分析病毒颗粒生物发生 联系人.这些技术在病毒学中具有广泛的适用性，这项研究的成功将是 适用于关于病毒复制和病毒-宿主细胞相互作用的各种问题。