Potassium Channel Trafficking in Geometrically Patterned Immunological Synapses

描述（由申请人提供）：淋巴细胞通过一系列事件对抗原作出反应，这些事件构成T细胞活化过程。当抗原是病原体或癌细胞时，T细胞激活是必要的和有益的，但当免疫系统识别出身体自身细胞的组成部分是外来的时，它就变得不受欢迎了。这种异常反应发生在自身免疫性疾病中。因此，研究正常和自身免疫T细胞的激活过程具有重要意义。T细胞的活化是通过T细胞与抗原呈递细胞（APC）的接触和免疫突触（is）的形成而启动的。IS是一个高度组织化的信号区，形成于T/APC界面，是激活响应充分发展所必需的。虽然IS形成过程的各个方面已经被彻底研究，但潜在的膜离子事件却知之甚少。Kv1.3钾通道在T细胞中表达，它们与T细胞受体复合物和IS的各种信号分子一起区隔。Kv1.3通道在T细胞活化过程中发挥重要作用，因为它们调节下游功能事件所需的Ca2+内流。事实上，抑制这些通道会终止免疫反应，因此Kv1.3阻滞剂正在作为一种新的免疫抑制剂被开发。尽管这些通道具有重要意义，但它们在T细胞IS中区隔化的功能后果尚未确定。我们实验室的初步数据表明，Kv1.3通道在IS内的位置对于调节通道功能是必要的，并且在自身免疫性疾病SLE中观察到Kv1.3通道定位的改变。在本应用中，我们将测试Kv1.3通道在免疫突触中的募集及其在该结构中的特定位置对于通道活性的调节是必要的，因此它影响T细胞激活的结果。具体来说，我们将设计人工apc样表面，允许在IS内预定位置靶向隔离Kv1.3蛋白，并同时测量下游功能事件。这些研究将确定工程蛋白表面用于研究IS中离子通道区隔化的可行性，特别是解决IS中离子通道位置如何决定T细胞/APC相互作用结果的问题。此外，这些研究将有助于理解在病理条件下，离子通道募集到IS中的异常对整体T细胞功能的影响。

DESCRIPTION (provided by applicant): Lymphocytes respond to antigens with a series of events that constitute the T cell activation process. T cell activation is necessary and beneficial when the antigen is a pathogen or a cancer cell, but it becomes undesirable when the immune system identifies components of the body's own cells as foreign. This aberrant response occurs in autoimmune diseases. Thus studying the activation process of normal and autoimmune T cells is of primary importance. T cell activation is initiated by contact of the T cell with the antigen presenting cell (APC) and formation of the immunological synapse (IS). The IS is a highly organized signaling zone that forms at the T/APC interface and is needed for full development of the activation response. Although various aspects of the process of IS formation have been thoroughly investigated, the underlying membrane ionic events are poorly understood. Kv1.3 potassium channels are expressed in T cells where they compartmentalize together with the T cell receptor complex and various signaling molecules at the IS. Kv1.3 channels play an important role during T cell activation as they regulate the Ca2+ influx necessary for downstream functional events. Indeed, inhibition of these channels terminates the immune response and therefore Kv1.3 blockers are under development as novel immunosuppressive agents. Despite the significance of these channels, the functional consequences of their compartmentalization in the IS of T cells are yet to be determined. Preliminary data from our laboratory suggest that Kv1.3 location within the IS is necessary to regulate the channel function and alterations in Kv1.3 channel localization in the IS have been observed in the autoimmune disease SLE. In this application we will be testing the hypothesis that Kv1.3 channels' recruitment in the immunological synapse and their specific location within this structure is necessary for regulation of the channel activity and consequently it influences the outcome of T cell activation. Specifically we will engineer artificial APC-like surfaces allowing targeted sequestration of Kv1.3 proteins in a predetermined location within the IS and simultaneous measurement of downstream functional events. These studies will establish the feasibility of engineered protein surfaces for studying ion channel compartmentalization in the IS, specifically addressing the question of how the position of ion channels in the IS determines the outcome of T cell/APC interactions. Furthermore, these studies will enable understanding of the implications that abnormalities in ion channel recruitment into the IS have on overall T cell function in pathological conditions. PUBLIC HEALTH RELEVANCE: Lymphocytes function, and also therefore malfunction, is in part controlled by Kv1.3 channels in the cell membrane. We are interested in studying details of how the location of Kv1.3 channels in the contact point between T cells and antigen presenting cells affects the channels' function. To do this we propose a novel method combining nanotechnology and artificial antigen presenting cell-like surfaces.