HTLV-1 Particle Analysis and Gag Interactions

描述(申请人提供)：人类T细胞白血病病毒(HTLV-1)感染全球约2000万人，是成人T细胞白血病/淋巴瘤(ATLL)的病原体，也可导致称为HTLV-1相关性脊髓病(HAM)/热带痉挛麻痹(TSP)的炎症性疾病综合征。在加勒比海盆地、中非和日本南部，普通人群中HTLV-1感染的流行率超过1%。HTLV-1以难以在细胞培养中研究而臭名昭著，这阻碍了对这些病毒如何在细胞中复制的严格分析，包括涉及逆转录病毒组装的步骤。即使对于其他更易处理的逆转录病毒系统，对逆转录病毒颗粒如何组装的细节也知之甚少。使用易于处理的模型系统、最先进的生物物理方法和一个跨学科的研究团队，我们进行了新颖的观察，形成了这一提议的基础。在这一应用中，我们建议使用多种实验方法来研究与HTLV-1颗粒大小、颗粒中的Gag化学计量以及活细胞中HTLV-1 Gag相互作用相关的问题。特别是，我们将应用冷冻电子显微镜/断层扫描(Cryo-EM/ET)、全内反射荧光(TIRF)显微镜和新颖的单分子荧光起伏光谱(FFS)技术来研究与以下问题有关的问题：1)颗粒尺寸和GAG化学计量；2)GAG靶向膜；以及3)HTLV-1颗粒的生物发生。这些拟议研究的结果应该会进一步深入了解HTLV-1和逆转录病毒颗粒组装的基本方面，这可能有助于开发治疗方法。 公共卫生相关性：人类T细胞白血病病毒1型(HTLV-1)是一种致癌的人类逆转录病毒，全世界约有2000万人感染，某些地区的流行率超过1%。对HTLV-1组装的基础研究将导致关于这些过程的详细信息，这将有助于更好地了解这些病毒如何在细胞中复制。这些信息可能会为新的治疗策略提供参考。

DESCRIPTION (provided by applicant): Human T-cell leukemia virus (HTLV-1) infects about 20 million individuals worldwide and is the etiological agent of an adult T-cell leukemia/lymphoma (ATLL), and can also result in an inflammatory disease syndrome called HTLV-1-associated myelopathy (HAM)/tropical spastic paraparesis (TSP). Prevalence rates for HTLV-1 infection in the general population are greater than 1% in the Caribbean Basin, Central Africa, and South Japan. HTLV-1 is notorious for being difficult to study in cell culture, which ha prohibited a rigorous analysis of how these viruses replicate in cells, including the steps involved in retrovirus assembly. The details for how retrovirus particle assembly occurs are poorly understood even for other more tractable retroviral systems. Using a tractable model system, state-of-the-art biophysical approaches, and an interdisciplinary research team, we have made novel observations that form the basis for this proposal. In this application, we propose to investigate questions related to HTLV-1 particle size, Gag stoichiometry in particles, and HTLV-1 Gag interactions in living cells using multiple experimental approaches. In particular, we will apply cryo-electron microscopy/tomography (cryo-EM/ET), total internal reflection fluorescence (TIRF) microscopy, and the novel single-molecule technology of fluorescence fluctuation spectroscopy (FFS) to investigate questions related to 1) particle size and Gag stoichiometry, 2) Gag targeting to membrane, and, 3) HTLV-1 particle biogenesis. The results from these proposed studies should provide further insight into fundamental aspects of HTLV-1 and retrovirus particle assembly, which may aid in developing therapeutics. PUBLIC HEALTH RELEVANCE: Human T-cell leukemia virus type 1 (HTLV-1) is a cancer-causing human retrovirus that infects about 20 million individuals worldwide, with prevalence rates greater than 1% in certain regions. Fundamental studies of HTLV-1 assembly will lead to detailed information about these processes that will be useful for a better understanding of how these viruses replicate in cells. Such information may inform new therapeutic strategies.