Immune regulation by SLAT, a Novel Th2-Expressed Protein

我们最近分离到了一种新的蛋白质，SLAT，它是RAC特异性激活子亚家族的成员。 SLAT在Th2细胞中上调，抗原刺激诱导其免疫突触定位和ZAP- 70关联，导致TCR招募和ZAP-70激活减少。我们两个，非排他性的。 工作假说是：1)SLAT通过其关联的ZAP-70抑制该激酶的TCR募集和/或 激活，从而以促进Th2分化的方式调节TCR信号的质量， 扩增和/或激活；以及2)SLAT和/或其替代产物SLAT2在Th2细胞中起作用 特定于CDC42/RAC的全环基金；这项活动在确定不同的 Th2细胞中IS和脂筏聚集的质量，以及由此产生的下游信号。我们将解决 并对SLAT的表达、功能和调控的其他方面进行了探讨 四个目标：1)使用原代小鼠T细胞，成像和生化技术，不同的刺激，以及 受体阻断抗体，我们将分析SLAT/SLAT2的mRNA或蛋白的表达，研究其 细胞内定位，MAP残基是膜/IS定位所必需的，并评估 这种本地化的意义。2)我们将对SLAT1小鼠进行详细的分析，包括Th1/Th2 分化、近端和下游信号事件、抗体和T细胞反应以及敏感性 与Th1或Th2表型占优势有关的实验性疾病。3)我们会 通过绘制SLAT和ZAP-70(或Syk)之间的图谱，探索SLAT和ZAP-70(或Syk)之间的生物学相关性 相互作用位点，并评估相互作用不足的SLAT或ZAP-70/Syk突变体或显性突变的影响 阴性的ZAP-70/Syk，对Th1/Th1的发育和激活。4)我们将使用生化和遗传技术 方法，包括SLAT1‘小鼠或用slat突变体重组的T细胞，以绘制slat残基的图谱 负责全球环境基金的活动，研究调节全球环境基金SLAT活动的机制，并确定 这一活动对SLAT的Th2偏斜效应和Th2中IS的独特组织的意义 细胞。这些研究将表征一种新的、多功能的TCR-近端蛋白。

We recently isolated a novel protein, SLAT, member of a novel subfamily of Rac-specific activators. SLAT is upregulated in Th2 cells and antigen stimulation induces its immune synapse localization and ZAP- 70 association, leading to reduced TCR recruitment and activation of ZAP-70. Our two, non-exclusive. working hypotheses are:1) SLAT, via its association ZAP-70, inhibits the kinase's TCR recruitment and/or activation, thereby modulating the quality of TCR signaling in a manner that promotes Th2 differentiation, expansion and/or activation; and 2) SLAT and/or its alternative product, SLAT2, functions in Th2 cells as a Cdc42/Rac-specific GEF; this activity plays a non-redundant and unique role in determining the distinct quality of the IS and lipid raft clustering and, consequently, downstream signals, in Th2 cells. We will address these hypotheses and analyze other aspects of the expression, function and regulation of SLAT by exploring four aims: 1) Using primary mouse T cells, imaging and biochemical techniques, different stimuli, and receptor-blocking antibodies, we will analyze the expression of SLAT/SLAT2 mRNA or protein, study their intracellular localization, map residues essential for the membrane/IS localization, and assess the significance of this localization. 2) We will conduct a detailed analysis of SLAT1" mice, including Th1/Th2 differentiation, proximal and downstream signaling events, antibody and T cell responses, and susceptibility to experimental diseases that are associated with predominance of the Th1 or Th2 phenotype. 3) We will explore the biological relevance of the association between SLAT and ZAP-70 (or Syk) by mapping the interaction sites, and assessing the effects of interaction-deficient SLAT or ZAP-70/Syk mutants, or dominant negative ZAP-70/Syk, on Th1/Th1 development and activation. 4) We will use biochemical and genetic approaches, including SLAT1' mice or T cells reconstituted with SLAT mutants, to map the SLAT residues responsible for GEF activity, study mechanisms that regulate the GEF activity of SLAT, and determine the significance of this activity for the Th2-skewing effect of SLAT and for distinct organization of the IS in Th2 cells. These studies will characterize a novel, multifunctional TCR-proximal protein.