Mechanisms of tunable posttranslational control of T-cell homeostasis and tolerance

摘要 在严格监管的过程中存在缺陷，这些过程产生并维持着多样化和自我容忍的 负责免疫的T细胞会导致非常虚弱的人类自身免疫和免疫缺陷疾病。 尽管经过几十年的发展，自身免疫性疾病仍然是导致死亡的主要原因之一。 特别是在妇女和儿童中，仅在美国就有2300多万人受到影响，代价是 每年为经济带来超过1000亿美元的收入。作为功能失调的T细胞发育和动态平衡 通常是自身免疫性疾病状态的基础，针对这些疾病的更有效治疗的发展 疾病将受益于解决细胞因子和途径的主要知识空白，使T- 细胞动态平衡。例如，依赖蛋白酶体的机制控制蛋白质的活性， 调节T细胞的存活、代谢和信号转导，但T细胞的关键动态平衡信号是如何 受体(TCR)和白介素7(IL-7)与泛素化机制的偶联仍然知之甚少。我们 最近发现，带电的多囊泡体蛋白-5(CHMP5)在 T细胞发展以招募促进客户蛋白质稳定性的去泛素酶。新证据来自 我们的实验室表明，CHMP5的表达受到TCR和IL-7信号的严格控制，并且 外周T细胞中CHMP5的缺失破坏了它们的动态平衡，并与全穿透性相关 多器官自身免疫性疾病。因此，利用允许精确追踪CHMP5的动物模型 突变的T细胞，在这个建议中，我们将检验CHMP5核关键的翻译后 内环境平衡信号与T细胞存活的蛋白质介质的稳定性相结合的节点 和耐受性，使其成为调节疾病中T细胞的一个可调和潜在的靶点。在AIM I中， 我们将确定CHMP5如何控制外周T细胞所必需的能量代谢和生存蛋白 生存和功能。在目标2中，我们将阐明TCR和TCR稳定CHMP5差异的机制 IL-7信号，特别是它们诱导丝氨酸磷酸化以稳定CHMP5蛋白的能力。 此外，由于脱泛素酶USP8的缺失会耗尽T细胞中的CHMP5蛋白，我们将定义 USP8-CHMP5相互作用的基础。破坏这种相互作用可能被用来治疗 耗尽T细胞CHMP5。到目前为止，TCR信号阈值是如何转化为胸腺细胞阳性和阴性的 具体的选择仍不清楚。因此，根据CHMP5由低亲和力TCR配体稳定的证据， 被高亲和力信号降解，在目标3中，我们将测试区分CHMP5蛋白的新范式 稳定是胸腺细胞建立中枢耐受的一种机制。这些研究将产生对 关于T细胞动态平衡的长期问题并有可能发现新的翻译后 可被利用来在治疗上调节疾病中T细胞的脆弱性，尤其是在调节失调时 在人类9p13.3染色体区域中，CHMP5基因的定位与多种组织的疾病相关。

Summary Defects in the stringently regulated processes that generate and maintain the diverse and self-tolerant pool of T-cells responsible for immunity cause very debilitating human autoimmune and immunodeficiency diseases. Despite decades of evolving therapies, autoimmune diseases still rank among the leading causes of death especially in women and children in the US alone where more than 23 million people are afflicted, at a cost that exceeds $100 billion annually to the economy. As dysfunctional T-cell development and homeostasis frequently underlie autoimmune disease states, development of more effective therapies against these diseases will benefit from resolving major knowledge gaps of the cellular factors and pathways that enable T- cell homeostasis. For example, proteasome-dependent mechanisms control the activity of proteins that mediate survival, metabolism and signal transduction in T-cells but how key homeostatic signals from the T-cell receptor (TCR) and interleukin (IL)-7 are coupled to the ubiquitination machinery is still poorly understood. We recently discovered that the Charged Multivesicular Body Protein-5 (CHMP5) functions as an “adaptor” during T-cell development to recruit deubiquitinating enzymes that promote client protein stability. New evidence from our laboratory shows that CHMP5 expression is stringently controlled by TCR and IL-7 signals, and that deletion of CHMP5 in peripheral T-cells impaired their homeostasis and was associated with a fully penetrant multi-organ autoimmune condition. Thus, leveraging animal models that allow precise tracking of CHMP5 mutant T-cells, in this proposal we will test the hypothesis that CHMP5 nucleates a critical posttranslational node by which homeostatic signals are integrated to the stability of protein mediators of T-cell survival and tolerance, situating it as a tunable and potential target for modulating T-cells in disease. In Aim I, we will determine how CHMP5 controls energy metabolism and prosurvival proteins integral to peripheral T-cell survival and function. In Aim 2, we will elucidate the mechanism of differential CHMP5 stabilization by TCR and IL-7 signals, especially their ability to induce serine phosphorylations that stabilize CHMP5 protein. Additionally, as deletion of the deubiquitinase USP8 depletes CHMP5 proteins in T-cells, we will define the basis of the USP8-CHMP5 interaction. Disrupting this interaction can potentially be utilized to therapeutically deplete T-cell CHMP5. To date, how TCR signal thresholds are translated into thymocyte positive and negative selection remains unclear. Thus, building on evidence that CHMP5 is stabilized by low affinity TCR ligands but degraded by high affinity signals, in Aim 3, we will test the novel paradigm that differential CHMP5 protein stabilization is a thymocyte mechanism to establish central tolerance. These studies will yield insights into long-standing questions on T-cell homeostasis and have the potential to uncover new posttranslational vulnerabilities that can be exploited to therapeutically modulate T-cells in disease especially as dysregulation of the human 9p13.3 chromosomal region location of CHMP5 gene is linked with diseases in multiple tissues.