Role of SHP-1 in T cell activation and development

描述(申请人提供)：SHP-1是一种蛋白酪氨酸磷酸酶，主要在造血细胞中表达，与细胞因子、生长因子和抗原诱导的信号事件的负调控有关。小鼠的SHP-1基因突变导致了蚊子(Me/Me)表型。Me等位基因纯合的小鼠，导致没有任何可检测到的SHP-1蛋白，在所有造血谱系中表现出各种疾病，导致出生后大约两到三周死亡。这些小鼠为将体外生化分析与体内和体外生物学研究相结合提供了宝贵的工具。我们的长期目标是了解SHP-1如何影响造血细胞的生长和分化。这项建议将试图阐明SHP-1参与了未成熟、成熟和调节T细胞的发育和功能，并结合了生物和生化方法。我们已经证明了SHP-1的一个亚群定位于脂筏，并且这种定位与TCR信号的功能相关。然而，靶向SHP-1的模式尚不清楚。目的1的目的是确定驱动SHP-1脂筏定位的分子机制以及脂筏定位的功能含义：我们的初步研究表明，SHP-1缺陷的小鼠胸腺和脾中的CD4+CD25+调节性T细胞数量增加。在目标2中，我们建议检验SHP-1影响CD4+CD25+Treg细胞功能的假设。我们将使用体外和体内实验检测SHP-1缺陷的Treg细胞的抑制潜力的强度和持续时间，以及SHP-1在Treg细胞内信号转导中的作用。在目标3中，我们将测试T细胞对SHP-1的内在需求，以及其他缺乏SHP-1活性的造血系在调节性T细胞群的生成/扩增中的作用。我们将使用携带转基因的小鼠来解决这些问题，以诱导显性负SHP-1突变体的诱导和组织特异性表达。综上所述，这里提出的研究应该会让我们更好地理解SHP-1和TCR信号。此外，我们希望更好地了解影响CD4+CD25+Treg细胞发育和功能的因素，重点是SHP-1在这些过程中的作用。

DESCRIPTION (provided by applicant): SHP-1 is a protein tyrosine phosphatase expressed predominantly in hematopoietic cells where it has been linked to negative regulation of signaling events induced by cytokines, growth factors and antigens. Mutations in the SHP-1 gene in mice cause the motheaten (me/me) phenotype. Mice homozygous for the me allele, which results in the absence of any detectable SHP-1 protein, display a variety of disorders in all hematopoietic lineages resulting in death about two to three weeks after birth. These mice provide a valuable tool to combine in vitro biochemical assays with in vivo and ex vivo biological studies. Our long term goal is to understand how SHP-1 influences the growth and differentiation of hematopoietic cells. This proposal will attempt to elucidate the involvement of SHP-1 in immature, mature and regulatory T cell development and function using a combination of biological and biochemical approaches. We have shown that a subpopulation of SHP-1 localizes to lipid rafts and that this localization is functionally relevant for TCR signaling. However, the mode of targeting SHP-1 is not known. The goal of Aim 1 is to determine the molecular mechanism driving lipid rafts localization of SHP-1 and the functional implications of lipid rafts localization: Our preliminary studies suggest that mice deficient in SHP-1 show increased numbers of CD4+CD25+ regulatory T cells in the thymus and spleen. In Aim 2, we propose to test the hypothesis that SHP-1 affects the function of CD4+CD25+ Treg cells. We will examine the strength and duration of the suppressive potential of SHP-1-deficient Treg cells using in vitro and in vivo assays, the role of SHP-1 in intracellular signaling of Treg cells. In Aim 3, we will test the T cell intrinsic requirement for SHP-1, and the role of other hematopoietic lineages deficient in SHP-1 activity on the generation/expansion of a regulatory T cell population. We will use mice carrying transgenes for inducible and tissue-specific expression of dominant negative SHP-1 mutants to address these questions. Taken together, the studies proposed here should give us a better understanding of the SHP-1 and TCR signaling. In addition, we expect to gain a better understanding of factors that influence the development and function of CD4+CD25+ Treg cells with a focus on the role of SHP-1 during these processes.