Regulation of Atopic Immune Responses by the Prolyl lsomerase Cyclophillin A

哮喘和其他过敏性疾病是工业化国家普遍存在的健康问题。为了 开发治疗药物以减轻这些疾病的严重性，这些疾病的生物过程 过敏症需要被定义。过敏反应是由辅助性T细胞对抗原的过度反应引发的； 因此，这些细胞是新疗法的诱人靶点。这项建议的长期目标是 确定辅助T细胞内控制过敏产生的特定细胞内信号事件- 促进IL-4等细胞因子的产生。我们已经确定了环孢素受体的重要作用。 亲环素A(CypA)在调节辅助T细胞反应中的作用CypA改变多肽键的结构 与脯氨酸残基相邻，因此可能通过诱导构象变化来调节蛋白质的功能。 缺乏CypA的小鼠会出现含有嗜酸性粒细胞和肥大细胞的炎症，这两种细胞类型具有 在过敏反应中的关键作用。这些小鼠的辅助性T细胞过度产生IL-4和其他过敏反应- 相关细胞因子并显示关键信号分子磷脂酶C-Gamma1的激活增加 (PLCgl)。已知的PLCgl调节因子是ITK，它是一种控制IL-4表达的酪氨酸激酶。CypA相互作用 与ITK中的脯氨酸残基；这种相互作用促进了ITK的自结合，这被认为是 下调ITK活性，并抑制ITK与其他促进PLCgl的因子之间的接触 激活。基于这些发现，我们的中心假设是CypA作为ITK活性的抑制因子， 从而限制辅助性T细胞表达IL-4。为了探索这一假设，我们将追求以下几点 具体目标：1)确定ITK与CypA相互作用和ITK自身所需的氨基酸 生化分析将被用来评估ITK和CypA中的氨基酸变化如何影响 蛋白质-蛋白质相互作用；2)确定CypA和ITK突变对功能的影响 功能改变的突变蛋白将在辅助T细胞中表达以评估其对IL-4的影响 表达和PLCgl激活；3)确定CypA和ITK在变态反应性疾病中的作用；如何改变 分析CypA或ITK活性在哮喘小鼠模型中的作用；4) 确定CypA是否调节表达过敏促进的辅助性T细胞的发展 细胞因子；细胞培养系统将被用来确定CypA的调控靶点。

Asthma and other allergic diseases are widespread health problems for industrialized nations. In order to develop therapeutic agents that lessen the severity of these diseases, the biological processes that drive allergy need to be defined. Allergic reactions are triggered by excessive helper T cell responses to antigens; thus these cells are attractive targets for new therapeutics. The long-term objective of this proposal is to define specific intracellular signaling events within helper T cells that control the production of allergy- promoting cytokines such as IL-4. We have identified an important role for the cyclosporine receptor cyclophilin A (CypA) in regulating helper T cell responses. CypA alters the structure of peptide bonds adjacent to proline residues, and may thus regulate protein function by inducing conformational changes. Mice lacking CypA develop inflammation that contains eosinophils and mast cells, two cell types that have key roles in allergic responses. Helper T cells from these mice overproduce IL-4 and other allergy- associated cytokines and show increased activation of the key signaling molecule phospholipase C-gamma1 (PLCgl). A known regulator of PLCgl is Itk,a tyrosine kinase that controls IL-4 expression. CypA interacts with a proline residue in Itk ; this interaction promotes Itk self-association, which is postulated to downregulate Itk activity, and also inhibits contacts between Itk and other factors that promote PLCgl activation. Based on these findings, our central hypothesis is that CypA functions as represser of Itk activity, thus limiting IL-4 expression by helper T cells. To explore this hypothesis, we will pursue the following specific aims: 1) Identify amino acids that are required for interaction between Itk and CypA and for Itk self- association; biochemical assays will be used to assess how amino acid changes in Itk and CypA affect protein-protein interactions; 2) Determine the effects of mutations in CypA and Itk on function; CypA and Itk mutant proteins with altered function will be expressed in helper T cells to assess their impact onIL-4 expression and PLCgl activation; 3) Determine the role of CypA and Itk in allergic disease; how changes in CypA or Itk activity modulate asthma development in mouse model for this disease will be analyzed; 4) Determine whether CypA regulates the development of helper T cells that express allergy-promoting cytokines; a cell culture system will be used to define regulatory targets for CypA.