Cytoskeletal regulation of T cell-APC interactions

描述（由申请人提供）：T细胞与抗原呈递细胞（APC）的相互作用在正常宿主免疫应答和包括自身免疫性疾病在内的病理条件下都很重要。尽管它们的重要性，我们仍然有有限的了解，调节T细胞-APC接触，T细胞活化的关键决定因素的形成和释放的机制。这项研究的长期目标是了解调节T细胞-APC接触持续时间的分子机制，以及这些3D粘附结构如何影响免疫反应和自身免疫性疾病的发展。虽然T细胞APC接触网站，称为免疫突触，共享许多结构组件与整合素介导的局灶性粘连，我们已经证明，调节T细胞APC接触网站动态的粘附机制是不同的。尽管最近取得了这一进展，但调节短暂与持续T细胞-APC接触形成以介导T细胞耐受或免疫的机制在很大程度上仍然未知。此外，组分进入和离开T细胞-APC接触位点的动力学尚未明确。指导我们研究的假设是粘着斑转换的关键调节因子talin、磷脂酰肌醇磷酸激酶PIPKI 390和粘着斑激酶（FAK）差异性地调节整联蛋白LFA-1的极化和T细胞- APC接触的动态组装和分解以影响T细胞活化和自身免疫。大量证据表明，稳定的T细胞-APC相互作用是T细胞活化和自身免疫性疾病发展所必需的。然而，介导持续的T细胞-APC相互作用的LFA-1的消耗可导致多发性硬化症（一种CD 4 T细胞驱动的疾病）的实验性自身免疫性脑炎（EAE）小鼠模型中自身免疫性恶化（Gultner，2010）。因此，粘附相互作用和自身免疫之间的关系可能难以预测。使用我们已经产生的基因敲除小鼠，我们将确定LFA-1相关蛋白，talin，PIPKI 390和FAK如何调节EAE中自身免疫的发展。我们现在处于独特的地位，可以实现以下具体目标：测试talin在T细胞-APC相互作用和T细胞活化中的作用。二.确定磷脂酰肌醇磷酸激酶（PIPKI 390）如何调节T细胞-APC相互作用和EAE中自身免疫的发展。三.测试粘着斑激酶（FAK）在T细胞- APC相互作用和T细胞活化中的作用。该提议的优势在于使用条件转基因小鼠，使用新型诱导试剂和实时3D延时荧光成像来探测调节T细胞-APC接触位点的动态以影响T细胞活化的途径。拟议的研究将导致更好地理解调节T细胞-APC相互作用和T细胞活化的机制，并可能为治疗自身免疫性疾病的新策略提供基础。

DESCRIPTION (provided by applicant): T cell interactions with antigen-presenting cells (APCs) are important both in normal host immune responses and in pathologic conditions including autoimmune disease. Despite their importance, we still have limited understanding of the mechanisms that regulate the formation and release of T cell-APC contacts, critical determinants of T cell activation. The long-term goal of the proposed research is to understand the molecular mechanisms that regulate the duration of T cell-APC contacts and how these 3D adhesive structures affect immune responses and the development of autoimmune disease. Although T cell-APC contact sites, known as the immune synapse, share many structural components with integrin-mediated focal adhesions, we have demonstrated that the adhesive mechanisms that regulate T cell-APC contact site dynamics are distinct. Despite this recent progress, the mechanisms that regulate the formation of transient versus sustained T cell-APC contacts to mediate T cell tolerance or immunity remain largely unknown. Moreover, the kinetics with which components enter and leave T cell-APC contact sites are not well defined. The hypothesis that guides our research is that key regulators of focal adhesion turnover, talin, phosphatidylinositide phosphate kinase PIPKI390 and focal adhesion kinase (FAK) differentially regulate the polarization of the integrin LFA-1 and the dynamic assembly and disassembly of T cell- APC contacts to affect T cell activation and autoimmunity. Substantial evidence suggests that stable T cell-APC interactions are necessary for T cell activation and the development of autoimmune disease. Yet, depletion of LFA-1, which mediates sustained T cell-APC interactions, can lead to worsening autoimmunity in the experimental autoimmune encephalitis (EAE) mouse model of multiple sclerosis, a CD4 T cell driven disease (Gultner, 2010). Therefore, the relationship between adhesive interactions and autoimmunity may be difficult to predict. Using knockout mice we have generated, we will determine how LFA-1 associated proteins, talin, PIPKI390 and FAK regulate the development of autoimmunity in EAE. We are now uniquely positioned to pursue the following specific aims: I. Test the role of talin in T cell-APC interactions and T cell activation. II. Determine how phosphatidylinositide phosphate kinase (PIPKI390) regulates T cell-APC interactions and the development of autoimmunity in EAE. III. Test the role of focal adhesion kinase (FAK) in T cell- APC interactions and T cell activation. The strength of this proposal lies in the use of conditional transgenic mice, use of novel inducible reagents and live 3D time-lapse fluorescent imaging to probe the pathways that regulate the dynamics of T cell-APC contact sites to affect T cell activation. The proposed studies will lead to improved understanding of the mechanisms that regulate T cell-APC interactions and T cell activation and may provide the foundation for new strategies to treat autoimmune disease.