ROLE OF FUCOSYLTRANSEFERASE IN ANTIVIRAL CYTOTOXICITY

岩藻糖基转移酶在抗病毒细胞毒性中的作用

• 批准号: 6031894
• 负责人: MANJUNATH NARASIMHA SWAMY
• 金额: $ 14.14万
• 依托单位: IMMUNE DISEASE INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-03-01 至 2001-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6031894
• 关键词: CD95 molecule; T cell receptor; antigen presentation; antigen presenting cell; cellular immunity; clinical research; cytotoxic T lymphocyte; enzyme activity; genetically modified animals; green fluorescent proteins; hexosyltransferase; human subject; immunodeficiency; interferon gamma; interleukin 2; interleukin 4; laboratory mouse; leukocyte activation /transformation; lymphocytic choriomeningitis virus; passive immunization; pore forming protein; selectins; tumor necrosis factor alpha

Cytotoxic T lymphocytes (CTL) are key immune effector cells in controlling viral infections and tumors. However, their differentiation in vivo is only partially understood. We have found that a novel APC-CD8 T cell interactions through an alpha (1,3) fucosylated P-selectin glycoprotein-1 (PSGL-1) is required to generate effector CTL. In its absence, vaccinia virus-infected mice deficient in alpha (1,3) fucosyl transferase IV+VIII (ft-/-) do not generate effector CTL, although they generate activated CD8 T cells capable of viral specific proliferation. In Aim 1, we will breed FT-/- mice to mice transgenic (Tg) to lymphocytic choriomeningitis (LCMV) glycoprotein TCR. This system will enable us to study the role of FT in antigen-specific CTL response that LCMV challenge. We will determine if FT is required on APC or on CD8 T cells in adoptive transfer experiments. Leukocyte adhesion deficiency type II (LADII), a human disease due to defective fucose metabolism allows us to study the role of fucosylated PSGL-1 in humans We have also produced transgenic mice whose T cells express the Jellyfish Victoria acquoria-derived green fluorescent protein (GFP-Tg). When these mice are immunized, the effector CTL lose GFP expression, whereas naive and activated pre-effector T cells continue to express GFP. Thus, GFP-Tg mice provide a powerful tool to separate differentiated effector from pre-effector CTL. Using this system, we have identified several previously unrecognized molecular changes that occur during effector differentiation. Thus, to study the molecular mechanism that are defective in the absence of T, in aim 2 we will breed FT-/- mice to GFP- Tg mice and analyze their antigen activated CD8 T cells for molecular differences from FT+/+ CD8 T cells. Since selectin-ligand deficient FT-/- mice, but not selectin-deficient mice are defective for CTL generation, a non-selectin receptor may exist for the fucosylated PSGL-1. Thus, in aim 3, we will determine if the defect in FT-/- mice is indirect through modulation of antigen presentation, or if a direct interaction with a novel molecule is involved. If such molecule exists, we will isolate, clone, and characterize the molecule.

细胞毒性T淋巴细胞（CTL）是控制病毒感染和肿瘤的关键免疫效应细胞。然而，它们在体内的分化仅被部分理解。我们已经发现一种新的APC-CD 8 T细胞相互作用通过α（1，3）岩藻糖基化的P-选择素糖蛋白-1（PSGL-1）是产生效应CTL所必需的。在其缺失的情况下，α（1，3）岩藻糖基转移酶IV+VIII（ft-/-）缺陷的牛痘病毒感染的小鼠不产生效应CTL，尽管它们产生能够进行病毒特异性增殖的活化的CD 8 T细胞。目的1：将FT-/-小鼠培育成淋巴细胞性脉络丛脑膜炎（LCMV）糖蛋白TCR转基因小鼠（Tg）。这一系统将使我们能够研究FT在LCMV攻击的抗原特异性CTL应答中的作用。我们将确定在过继转移实验中是否需要在APC或CD 8 T细胞上进行FT。白细胞粘附缺陷II型（LADII），一种由于岩藻糖代谢缺陷引起的人类疾病，使我们能够研究岩藻糖基化PSGL-1在人类中的作用。我们还产生了转基因小鼠，其T细胞表达水母维多利亚水母衍生的绿色荧光蛋白（GFP-Tg）。当这些小鼠被免疫时，效应CTL失去GFP表达，而初始和活化的前效应T细胞继续表达GFP。因此，GFP-Tg小鼠提供了一个强大的工具，以区分分化的效应子从前效应子CTL。使用这个系统，我们已经确定了一些以前未被认识到的分子变化，发生在效应分化。因此，为了研究在T不存在下有缺陷的分子机制，在目标2中，我们将FT-/-小鼠培育成GFP-Tg小鼠，并分析它们的抗原活化的CD 8 T细胞与FT+/+ CD 8 T细胞的分子差异。由于选择素配体缺陷型FT-/-小鼠而非选择素缺陷型小鼠在CTL生成方面存在缺陷，因此岩藻糖基化PSGL-1可能存在非选择素受体。因此，在目标3中，我们将确定FT-/-小鼠中的缺陷是否是通过调节抗原递呈间接产生的，或者是否涉及与新分子的直接相互作用。如果这样的分子存在，我们将分离，克隆和表征分子。