TRAIL receptor in apoptosis and the immune system

TRAIL受体在细胞凋亡和免疫系统中的作用

• 批准号: 6359738
• 负责人: ASTAR WINOTO
• 金额: $ 29.54万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6359738
• 关键词: RNA splicing; Retroviridae; apoptosis; complementary DNA; cytokine receptors; gene mutation; gene targeting; genetically modified animals; laboratory mouse; lymphocyte; receptor expression; transfection /expression vector; tumor necrosis factor alpha

DESCRIPTION: (provided by applicant): TRAIL (tumor necrosis factor-related apoptosis inducing ligand) is a member of the tumor necrosis factor superfamily. It can kill a variety of tumor cells but has no appreciable effect on most normal cells. Several TRAIL receptors have been identified in human. These include DR4, DR5, DcR1 and DcR2. DR4 and DR5 contain a death domain in their cytoplasmic tails and can initiate cell death when over-expressed. In contrast, DcRl and DcR2 do not encode any death domain and cannot initiate apoptosis. These so-called decoy receptors, DcR1 and DcR2, can instead act as a dominant negative mutant for DR4 and DR5, at least in transient transfection experiments. The physiological function of TRAIL and its receptors are poorly understood. Recently, soluble human DRS protein was shown to exacerbate collagen-induced autoimmunity and exogenous TRAIL was shown to inhibit T cell proliferation. Thus, TRAIL might act as a negative regulatory molecule for the immune system. To understand the physiological function of TRAIL and its receptor(s), a mouse cDNA clone that encodes a protein with equal homology to human DR4 and DR5 was isolated (rnDR4/5). An alternative splicing of the same gene was found to yield an identical protein without a death domain (mDcR). Transient transfection of mDR4I5 but not mDcR leads to apoptosis, and stable expression of mDR4/5 in FADD+ cells confers TRAIL sensitivity. To elucidate the role of this TRAIL receptor and its cytoplasmic-truncated form (mDcR) in the immune system, mice lacking both proteins and mice lacking DcR will be generated. Tissue-specific null mDR4/5 will also be made using the cre-loxP technology. The effect of DR4/5 and DcR mutation on lymphocyte development, T and B cell tolerance and immune responses will be examined. Similar to FasL, TRAIL-mediated apoptosis is mediated through FADD. This raises the question to how the same cells can be resistant to TRAIL but are sensitive to FasLmediated apoptosis. A receptor cytoplasmic-tail swapping experiment to dissect the underlying reasons for this is proposed. Retroviruses encoding mDR4/5-Fas (DR4/5 extracellular domain fused to the Fas death domain) or the reciprocal Fas-mDR4/5 chimeric proteins will be used to infect wild type or lpr/lpr T cells. The apoptotic effect of TRAIL or FasL on T cells expressing these chimeric proteins will be assessed. Successful completion of these experiments should lead to a significant understanding of how cancer differs from normal cells and how TRAIL receptor functions in the immune system.

描述：（申请人提供）：TRAIL（肿瘤坏死因子相关 凋亡诱导配体）是肿瘤坏死因子的成员 超家族。它可以杀死各种肿瘤细胞，但没有明显的作用 在大多数正常细胞上。在人类中已经确定了几个越野径受体。 其中包括DR4，DR5，DCR1和DCR2。 DR4和DR5在 它们的细胞质尾巴，可以在表达过表达时引发细胞死亡。在 对比，DCRL和DCR2不编码任何死亡领域，也无法启动 凋亡。这些所谓的诱饵受体DCR1和DCR2可以充当 DR4和DR5的主要负突变体，至少在瞬时转染中 实验。步道及其受体的生理功能很差 理解。最近，可溶性人DRS蛋白被证明加剧了 胶原蛋白引起的自身免疫性和外源性步道已显示可抑制T细胞 增殖。因此，TRAIL可能充当负调节分子 免疫系统。了解步道的生理功能及 受体，一种小鼠cDNA克隆，该克隆编码具有相同同源性的蛋白质 人DR4和DR5分离出来（RNDR4/5）。相同的替代剪接 发现基因产生没有死亡结构域（MDCR）的相同蛋白质。 MDR4I5但MDCR的瞬时转染会导致凋亡，并且稳定 MDR4/5在FADD+细胞中的表达赋予跟踪灵敏度。阐明 该路径受体的作用及其细胞质截断形式（MDCR）在 免疫系统，缺乏蛋白质的小鼠和缺乏DCR的小鼠将是 生成。还将使用Cre-loxP制造组织特异性的零MDR4/5 技术。 DR4/5和DCR突变对淋巴细胞发育的影响，T 将检查B细胞的耐受性和免疫反应。类似于fasl， 通过FADD介导了径向介导的细胞凋亡。这将问题提高到 相同的单元如何抗性，但对Faslsived敏感 凋亡。受体细胞质尾交换实验，以剖析 提出了基本原因。编码MDR4/5-FAS的逆转录病毒 （DR4/5的细胞外域融合到FAS死亡领域）或倒数 FAS-MDR4/5嵌合蛋白将用于感染野生型或LPR/LPR T 细胞。 TRAIL或FASL对表达T细胞的凋亡作用 将评估嵌合蛋白。这些实验成功完成 应该使人们对癌症与正常情况有所不同有深入的了解 细胞以及TRAIN受体如何在免疫系统中起作用。