Asb2 in CD4+ T cell lineage differentiation and its plasticity

描述（由申请人提供）：该R21申请的总体目标是探索含锚定重复SOCS盒蛋白2（Asb 2）在调节CD 4辅助T细胞分化中的作用。我们以前的研究表明，Asb 2基因是转录激活的Notch信号和Asb 2介导的Notch诱导的蛋白泛素化，通过桥接形成的二聚体E3泛素连接酶复合物。Asb 2促进大量蛋白质的降解，包括一些与T辅助细胞分化相关的蛋白质。有趣的是，全基因组研究表明，Asb 2在Th 2细胞中高度表达，在Treg细胞中受到抑制。有趣的表达模式表明Asb 2在辅助性T细胞分化中的潜在作用。我们假设：（1）Asb 2对T2分化和维持Th 2谱系命运至关重要;（2）Asb 2有利于T效应细胞命运，同时抑制Treg分化和破坏Treg鉴定。我们已经产生了小鼠模型，其中Asb 2功能的获得或丧失可以通过Cre介导的重组实现。这些动物模型将用于测试这些假设在两个目标。Aim 1将探索Asb 2在Th 2分化中的作用和Th 2命运的稳定性。我们将使用 Asb 2敲除小鼠，以测试Asb 2的缺失是否会损害体外Th 2分化或使细胞不稳定。 当切换到Th 1、Th 17或Treg的极化条件时，Th 2的命运。然后，我们将评估Asb 2在Th 2应答中的作用，通过用寄生虫抗原、来自曼氏血吸虫的可溶性卵抗原（SEA）或用明矾中的OVA免疫野生型和Asb 2-/-小鼠。将测量免疫小鼠中的Th 2细胞因子分泌和Th 2依赖性IG产生。Aim 2将探索Asb 2在抑制Treg分化中的作用。我们将在初始T细胞中从ROSA 26启动子异位表达Asb 2，并测试Asb 2是否可以在体外阻断iTreg分化，或者Asb 2是否可以增强Treg细胞向Th 1，Th 2和Th 17谱系的转化。我们还将在Asb 2表达小鼠体内检查Treg分化，并评估Asb 2表达骨髓拯救Foxp 3缺陷型皮屑骨髓移植到RAG 1缺陷型宿主中引起的免疫疾病的能力。这些研究结果将为Asb 2在辅助性T细胞分化中的作用提供初步评估，并为全面深入的研究奠定基础。这条调查路线审查了 辅助性T细胞分化的独特视角，即泛素介导的降解调节因子的分化。这些研究可能会产生新的想法，在免疫和自身免疫的竞争中具有治疗价值。

DESCRIPTION (provided by applicant): The overall objective of this R21 application is to explore the role of ankyrin-repeat SOCS-box containing protein 2 (Asb2) in the regulation of CD4 helper T cell differentiation. Our previous studies have shown that the Asb2 gene is transcriptionally activated by Notch signaling and Asb2 mediates Notch-induced protein ubiquitination through bridging the formation of dimeric E3 ubiquitin ligase complexes. Asb2 promotes the degradation a large number of proteins including some related to T helper cell differentiation. Interestingly, genome-wide studies have revealed that Asb2 is highly expressed in Th2 cells and repressed in Treg cells. The interesting pattern of expression suggests a potential role for Asb2 in helper T cell differentiation. We hypothesize: (1) Asb2 is crucial for T2 differentiation and maintenance of Th2 lineage fate; (2) Asb2 favors T effector cell fates while suppressing Treg differentiation and destabilizing Treg identifies. We have already generated mouse models in which gain or loss of Asb2 function can be achieved via Cre- mediated recombination. These animal models will be used to test these hypotheses in two aims. Aim1 will explore the role of Asb2 in Th2 differentiation and the stability of the Th2 fate. We will use Asb2 knockout mice to test if loss of Asb2 impairs Th2 differentiation in vitro or destabilizes the Th2 fate when switched to polarizing conditions for Th1, Th17 or Treg. We will then evaluate the role of Asb2 in Th2 responses in vivo by immunizing wild type and Asb2-/- mice with a parasite antigen, the soluble egg antigen (SEA) from Schistosoma mansoni or with OVA in alum. Th2 cytokine secretion and Th2-dependent Ig production in immunized mice will be measured. Aim2 will explore the role of Asb2 in suppressing Treg differentiation. We will ectopically express Asb2 from the ROSA26 promoter in na¿ve T cells and test if Asb2 can block iTreg differentiation in vitro or if Asb2 potentiates the conversion of Treg cells into Th1, Th2 and Th17 lineages. We will also examine Treg differentiation in vivo in Asb2-expressing mice and assess the ability of Asb2- expressing bone marrow to rescue the immune disorder caused by Foxp3-deficient scurfy bone marrow transplanted into RAG1 deficient hosts. Finding from these studies will provide an initial evaluation concerning the role of Asb2 in T helper cell differentiation and lay the ground work for a full-blown in-depth investigation. This line of investigation examines the regulation of T helper cell differentiation from a unique perspective, namely ubiquitin-mediated degradation of regulators of the differentiation. Fresh ideas will likely emerge from these studies that will be o therapeutic value in the contest of immunity and autoimmunity.