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Lipid rafts, exosomes, and human T-cell leukemia virus type 1 biology: A new model of viral pathogenesis.

机译：脂质筏，外来体和人类T细胞白血病病毒1型生物学：病毒发病机制的新模型。

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摘要

Previous studies in the Hildreth laboratory determined that the expression of VCAM-1 in the erythroleukemic cell line K562 caused susceptibility to HTL-V-1-induced syncytium formation. The observation that not all cells expressing VCAM-1 are sensitive to syncytium formation by the virus indicated that K562 cells express a second molecule which, in the presence of VCAM-1, renders the cells susceptible to syncytium formation. Immunization of mice with K562 cells and generation of hybridomas resulted in four monoclonal antibodies that blocked HTLV-1-induced syncytium formation between K562/VCAM1 cells and HTLV-1-infected MT2 cells. Cell phenotyping and immunoblotting indicated that all four MAbs recognized distinct proteins. Treatment of cells with phosphatidylinositol-specific phospholipase C determined that three of the four MAbs recognized GPI-anchored proteins, which are enriched in lipid raft regions of the plasma membrane. Lipid raft isolation and immunoblot analysis determined that proteins recognized by all four MAbs localized largely if not entirely to lipid rafts. Dispersion of lipid rafts by cholesterol depletion inhibited HTLV-1-induced syncytium formation without affecting the ability of VCAM-1 to bind its ligand, indicating the importance of lipid rafts in HTLV-1 biology. One of the four MAbs, K5.M1, whose target protein was deemed a particularly strong candidate as a virus receptor, recognized a 17 kD protein. Through affinity purification, this protein was isolated, sequenced by tandem mass spectrometry, and identified as Leu-13 antigen. Lipid raft marker CD59 was found on the viral envelope as determined by virus capture assays, and we investigated whether HTLV-1 used lipid rafts as sites for budding. Immunoblot, virus capture, and confocal microscopy data indicated that proteins on HTLV-1 virions are found on lipid raft domains in infected cells. However, some proteins which were in lipid rafts were not found on the virus. This observation, in addition to the high levels of MHC class II and tetraspanner protein incorporation into the virions led us to examine the possibility of a subcellular origin of the virions. The data presented in this study indicates that HTLV-1 virions carry many traits associated with exosomes.
机译：Hildreth实验室的先前研究确定，VCAM-1在红白血病细胞系K562中的表达引起对HTL-V-1诱导合胞体形成的敏感性。并非所有表达VCAM-1的细胞都对病毒对合胞体形成敏感的观察结果表明，K562细胞表达第二个分子，在VCAM-1存在下，第二个分子使细胞易于形成合胞体。用K562细胞免疫小鼠并产生杂交瘤，产生了四种单克隆抗体，它们阻断了HTLV-1诱导的K562 / VCAM1细胞与HTLV-1感染的MT2细胞之间的合胞体形成。细胞表型和免疫印迹表明，所有四个单克隆抗体均识别不同的蛋白质。用磷脂酰肌醇特异性磷脂酶C处理细胞后，确定四个MAb中的三个可识别GPI锚定的蛋白，这些蛋白富含在质膜的脂筏区域。脂质筏的分离和免疫印迹分析确定了所有四个单克隆抗体识别的蛋白质大部分（如果不是全部）定位于脂质筏。胆固醇耗尽对脂筏的分散抑制了HTLV-1诱导的合胞体的形成，而不影响VCAM-1结合其配体的能力，这表明脂筏在HTLV-1生物学中的重要性。四个单克隆抗体之一K5.M1（其目标蛋白被认为是作为病毒受体的特别强的候选物）识别出17 kD蛋白。通过亲和纯化，分离出该蛋白，通过串联质谱法测序，并鉴定为Leu-13抗原。脂质筏标记CD59被发现在病毒的包膜上，通过病毒捕获分析确定，我们调查了HTLV-1是否使用脂质筏作为出芽的位点。免疫印迹，病毒捕获和共聚焦显微镜数据表明，HTLV-1病毒体上的蛋白质在感染细胞的脂质筏结构域上发现。但是，在病毒中未发现脂质筏中的某些蛋白质。除了高水平的II类MHC和四跨膜蛋白掺入病毒粒子外，这一观察结果还使我们研究了病毒粒子亚细胞起源的可能性。这项研究中提供的数据表明，HTLV-1病毒体具有许多与外泌体相关的特征。

著录项

  • 作者

    Niyogi, Kakoli.;

  • 作者单位

    The Johns Hopkins University.;

  • 授予单位 The Johns Hopkins University.;
  • 学科 Biology Microbiology.
  • 学位 Ph.D.
  • 年度 2003
  • 页码 149 p.
  • 总页数 149
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 微生物学;
  • 关键词

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