Understanding how TRAF6 controls T lymphocyte activation

了解 TRAF6 如何控制 T 淋巴细胞激活

• 批准号: 1913536
• 金额: --
• 依托单位: University of Dundee
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2017
• 资助国家: 英国
• 起止时间: 2017 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-1913536
• 关键词: Understanding; how; TRAF6; controls; lymphocyte

The expression of TRAF6 (TNF Receptor-Associated Factor 6) is essential for many biological processes, which include signaling networks of the innate immune system, the production of osteoclasts and bone formation, lymph node organogenesis, and the development of hair follicles, sweat and sebaceous glands. TRAF6 is also needed for dendritic cell, B cell and T-cell function. It is a tenet of the field of innate immunity that the essential role of TRAF6, an E3 ubiquitin ligase activity, is to generate Lys63-linked ubiquitin chains, which then activate TAK1, the "master kinase" of the system. Recently, one of our laboratories (PC) found that this long-held view is incorrect, since innate immune signaling could be restored by the re-expression of E3 ligase-inactive mutants of TRAF6 in TRAF6 knock-out (KO) cells. These studies also revealed that the E3 ligase activity of TRAF6 was not essential because two other E3 ligases, Pellino1 and Pellino2, are also able to generate the Lys63-linked ubiquitin chains required for signaling. An important conclusion from these studies was that the essential roles of TRAF6 in innate immunity are independent of its E3 ligase activity (1). The PC lab also generated knock-in mice in which TRAF6 was replaced by the E3 ligase-inactive TRAF6[L74H] mutant. In contrast, to TRAF6 KO mice that have deformed bones and no teeth, the TRAF6[L74H] mice have normal bones and teeth, because the RANKL signaling pathway required for osteoclast formation is unimpaired in TRAF6[L74H] cells (1). These remarkable findings have raised the question of what the physiological role of the TRAF6 E3 ligase might be? Previous studies showed that TRAF6 has a crucial role to restrain T cell effector function (2). Hence T cell selective deletion of TRAF6 results in deregulation of T cell function and the development of T cell mediated autoimmunity. Analysis of TRAF6[L74H] mice by the PC laboratory indicates that they have hyperactive T cells in lymph nodes and spleens. Moreover, these mice develop autoimmune inflammatory diseases indicating that the E3 ubiquitin ligase activity of TRAF6 has a critical role in T cells to restraint cell effector function. Since DC's lab is pre-eminent in the field of T cell signaling, the PC and DC labs propose to jointly supervise the PhD student who will elucidate the molecular mechanism by which TRAF6 restricts T cell activation. The first part of the project will explore the development of effector and regulatory T cells in the secondary lymphoid tissues, gut and lung of TRAF6[L74H] mice. It will also explore the ability of TRAF6[L74H] T cells to mediate in vivo immune responses to pathogens and cancer and their role in autoimmunity. The second part will investigate the mechanisms by which TRAF6 activation suppresses T cell function which, in principle, could occur at any step in T cell signaling pathways. Pellino isoforms may function redundantly with TRAF6 and the project may benefit from the availability of Pellino knock-in mice developed in PC's laboratory, in which all three Pellinos have been replaced by E3 ligase-inactive mutants, alone and in combination. It has been described that the TRAF6 expression is required for IL-18 signaling in CD4 T cells and inhibits TGF-mediated suppression of IL-2 (3). We will therefore focus initially on how TRAF6 contributes to the signalling pathways used by these cytokines to control T cell biology. A fundamental strategy will be to use high resolution mass spectrometry to precisely map how IL-18 regulates the proteome, phosphoproteome and ubiquitome of wild type and TRAF6[L74H] CD4 T cells. How the signaling pathways controlled by these cytokines are integrated to control the cellular response of CD4 T cells is poorly understood at the molecular level and the complementary expertise of the DC and PC labs may enable important aspects of this problem to be cracked.

TRAF6（TNF 受体相关因子 6）的表达对于许多生物过程至关重要，包括先天免疫系统的信号网络、破骨细胞的产生和骨形成、淋巴结器官发生以及毛囊、汗液和皮脂腺的发育。 TRAF6 也是树突状细胞、B 细胞和 T 细胞功能所必需的。先天免疫领域的一个原则是，TRAF6（一种 E3 泛素连接酶活性）的基本作用是生成 Lys63 连接的泛素链，然后激活系统的“主激酶”TAK1。最近，我们的一个实验室 (PC) 发现这种长期持有的观点是不正确的，因为先天免疫信号可以通过在 TRAF6 敲除 (KO) 细胞中重新表达 TRAF6 E3 连接酶失活突变体来恢复。这些研究还表明，TRAF6 的 E3 连接酶活性并不是必需的，因为另外两种 E3 连接酶 Pellino1 和 Pellino2 也能够生成信号传导所需的 Lys63 连接的泛素链。这些研究的一个重要结论是 TRAF6 在先天免疫中的重要作用与其 E3 连接酶活性无关 (1)。 PC 实验室还生成了敲入小鼠，其中 TRAF6 被 E3 连接酶失活的 TRAF6[L74H] 突变体取代。相比之下，与骨骼变形且没有牙齿的 TRAF6 KO 小鼠相比，TRAF6[L74H] 小鼠具有正常的骨骼和牙齿，因为破骨细胞形成所需的 RANKL 信号通路在 TRAF6[L74H] 细胞中未受损 (1)。这些显着的发现提出了一个问题：TRAF6 E3 连接酶的生理作用可能是什么？先前的研究表明 TRAF6 对抑制 T 细胞效应功能具有至关重要的作用 (2)。因此，T 细胞选择性删除 TRAF6 会导致 T 细胞功能失调和 T 细胞介导的自身免疫的发展。 PC 实验室对 TRAF6[L74H] 小鼠的分析表明，它们的淋巴结和脾脏中存在高度活跃的 T 细胞。此外，这些小鼠出现自身免疫性炎症疾病，表明TRAF6的E3泛素连接酶活性在T细胞中对抑制细胞效应功能具有关键作用。由于 DC 实验室在 T 细胞信号转导领域处于领先地位，PC 和 DC 实验室提议共同指导博士生，他将阐明 TRAF6 限制 T 细胞激活的分子机制。该项目的第一部分将探索 TRAF6[L74H] 小鼠次级淋巴组织、肠道和肺中效应和调节性 T 细胞的发育。它还将探索 TRAF6[L74H] T 细胞介导体内针对病原体和癌症的免疫反应的能力及其在自身免疫中的作用。第二部分将研究 TRAF6 激活抑制 T 细胞功能的机制，原则上，这种机制可能发生在 T 细胞信号传导途径的任何步骤。 Pellino 亚型可能与 TRAF6 一起发挥冗余功能，该项目可能会受益于 PC 实验室开发的 Pellino 敲入小鼠的可用性，其中所有三种 Pellino 均已被单独或组合的 E3 连接酶失活突变体取代。据描述，TRAF6 表达是 CD4 T 细胞中 IL-18 信号转导所必需的，并且会抑制 TGF 介导的 IL-2 抑制 (3)。因此，我们首先将重点关注 TRAF6 如何促进这些细胞因子用于控制 T 细胞生物学的信号通路。一个基本策略是使用高分辨率质谱精确绘制 IL-18 如何调节野生型和 TRAF6[L74H] CD4 T 细胞的蛋白质组、磷酸化蛋白质组和泛素组。在分子水平上，人们对这些细胞因子控制的信号通路如何整合以控制 CD4 T 细胞的细胞反应知之甚少，DC 和 PC 实验室的互补专业知识可能有助于解决这一问题的重要方面。