Regulation of T-cell function and autoimmune inflammation by deubiquitinases.

 描述（由申请人提供）：泛素化是调节T细胞活化和T细胞介导的免疫应答的关键机制，并且失调的泛素化事件与包括自身免疫和炎症在内的人类免疫性疾病相关。泛素化的一个众所周知的功能是通过介导CD 3和关键信号因子的泛素依赖性降解来控制T细胞受体（TCR）近端信号事件。泛素化还以正或负方式调节TCR途径中下游信号传导分子的命运或活化。与磷酸化一样，泛素化是一个可逆的过程，其中泛素链分别被泛素化酶（E1，E2，E3）和去泛素化酶（DUBs）缀合和解缀合。尽管对E3泛素连接酶进行了广泛的研究，但调节TCR信号传导和T细胞功能的DUB仍然定义不清。PI的实验室在这一研究领域一直处于领先地位。在之前的资助周期中，我们已经取得了开创性的发现，证明了DUB CYLD在调节T细胞活化和自身免疫性炎症中的关键作用。此外，我们最近发表的工作和初步研究已经鉴定了几种新型T细胞调节DUB，这为这种更新申请奠定了坚实的基础。使用生物化学和小鼠遗传方法，我们已经将Otud 7 b鉴定为正调节TCR近端信号传导的DUB。Otud 7 b缺陷减弱TCR近端激酶Zap 70以及多种下游信号传导因子的活化。有趣的是，在对TCR/CD 28刺激的反应中，Otud 7 b被迅速募集到CD 3？，并且似乎调节该TCR的稳定性。 信号链基于这些发现，我们假设Otud 7 b可能通过调节CD 3？和可能的关键TCR近端信号传导因子的泛素化来促进TCR信号传导。我们的研究还鉴定了USP家族的两个同源DUB，Usp 4和Usp 15，作为T细胞活化的负调节剂。有趣的是，尽管Usp 4和Usp 15具有很强的同源性，但它们中任何一种的基因消融都会导致T细胞的过度活化。这一发现提出了一个问题，这两个高度同源的DUB如何发挥非冗余信号功能，以及它们是否在某些信号功能中也具有冗余。本继续申请的总体目标是了解Otud 7 b和Usp 4/Usp 15调节TCR信号传导和T细胞介导的免疫和自身免疫性炎症的分子机制。为了实现我们的总体目标，我们将（1）确定Otud 7 b介导TCR信号传导的分子机制，（2）研究Usp 4和USP 15如何负调节TCR信号传导，以及（3）研究Otud 7 b和Usp 4在调节T细胞功能和自身免疫性炎症中的体内功能。我们相信，这些拟议的研究具有高度创新性，将产生高影响力的发现，导致该领域的重大进步。随着新生成的小鼠模型的建立和我们丰富的经验，我们处于一个独特的位置来执行拟议的项目。

 DESCRIPTION (provided by applicant): Ubiquitination is a crucial mechanism that regulates T-cell activation and T cell-mediated immune responses, and deregulated ubiquitination events are linked to human immunological disorders including autoimmunity and inflammation. A well-known function of ubiquitination is to control T cell receptor (TCR)-proximal signaling events by mediating ubiquitin-dependent degradation of CD3¿ and key signaling factors. Ubiquitination also regulates the fate or activation of downstream signaling molecules in the TCR pathway, in a positive or negative manner. Like phosphorylation, ubiquitination is a reversible process, in which ubiquitin chains are conjugated and deconjugated by ubiquitinating enzymes (E1, E2, E3) and deubiquitinases (DUBs), respectively. Despite the extensive studies of E3 ubiquitin ligases, the DUBs that regulate TCR signaling and T-cell functions are still poorly defined. The PI's laboratory has been in a leading position in this research area. During the previous funding cycles, we have made seminal findings demonstrating a crucial role for the DUB CYLD in regulating T-cell activation and autoimmune inflammation. Moreover, our recently published work and preliminary studies have led to the identification of several novel T cell-regulatory DUBs, which forms a solid foundation for this renewal application. Using biochemical and mouse genetic approaches, we have identified Otud7b as a DUB that positively regulates TCR-proximal signaling. Otud7b deficiency attenuates the activation of the TCR-proximal kinase Zap70 as well as multiple downstream signaling factors. Interestingly, in response to TCR/CD28 stimulation, Otud7b is rapidly recruited to CD3¿ and appears to regulate the stability of this TCR signaling chain. Based on these findings, we hypothesize that Otud7b may facilitate TCR signaling by regulating the ubiquitination of CD3¿ and possibly key TCR-proximal signaling factors. Our studies also led to the identification of two homologous DUBs of the USP family, Usp4 and Usp15, as negative regulators of T-cell activation. Interestingly, despite the strong homology of Usp4 and Usp15, genetic ablation of either of them causes hyper-activation of T cells. This finding raises the question of how these two highly homologous DUBs exert non-redundant signaling functions and whether they also have redundancies in some signaling functions. The overall objective of this continuation application is to understand the molecular mechanisms by which Otud7b and Usp4/Usp15 regulate TCR signaling and T cell-mediated immunity and autoimmune inflammation. To accomplish our overall objective, we will (1) define the molecular mechanism by which Otud7b mediates TCR signaling, (2) examine how Usp4 and USP15 negatively regulate TCR signaling, and (3) investigate in vivo functions of Otud7b and Usp4 in regulating T-cell function and autoimmune inflammation. We believe that these proposed studies are highly innovative and will yield high-impact findings that lead to a major advancement of the field. With the establishment of newly generated mouse models and our extensive experience, we are in a unique position to carry out the proposed project.