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做出反应，这些事件构成了T细胞的激活过程。当抗原是病原体或癌细胞时，T细胞激活是必要的，也是有益的，但当免疫系统发现人体自身细胞的成分是外来时，激活T细胞就变得不可取了。这种异常反应发生在自身免疫性疾病中。因此，研究正常T细胞和自身免疫T细胞的激活过程具有重要意义。T细胞与抗原提呈细胞(APC)的接触和免疫突触(IS)的形成启动了T细胞的激活。IS是在T/APC界面形成的高度组织的信号区，是充分发展激活反应所必需的。虽然已经对IS形成过程的各个方面进行了彻底的研究，但对潜在的膜离子事件了解很少。Kv1.3钾通道在T细胞中表达，它们与T细胞受体复合体和IS处的各种信号分子一起分隔。Kv1.3通道在T细胞激活过程中起着重要作用，因为它们调节下游功能事件所需的钙离子内流。事实上，抑制这些通道会终止免疫反应，因此Kv1.3阻滞剂正在作为新的免疫抑制剂开发。尽管这些通道具有重要意义，但它们在T细胞内区隔的功能后果仍有待确定。我们实验室的初步数据表明，Kv1.3在IS中的位置是调节通道功能所必需的，并且在自身免疫性疾病SLE中已经观察到Kv1.3在IS中的通道定位的改变。在这一应用中，我们将检验一种假设，即Kv1.3通道在免疫突触中的招募及其在该结构中的特定位置对于调节通道的活性是必要的，因此它影响T细胞激活的结果。具体地说，我们将设计人工APC样表面，允许Kv1.3蛋白在IS内的预定位置进行定向隔离，并同时测量下游功能事件。这些研究将建立用于研究IS中离子通道区隔的工程蛋白质表面的可行性，特别是解决IS中离子通道的位置如何决定T细胞/APC相互作用的结果的问题。此外，这些研究将使人们能够理解离子通道募集到IS中的异常对病理条件下整体T细胞功能的影响。 与公共卫生相关：淋巴细胞的功能，以及由此产生的功能障碍，在一定程度上是由细胞膜上的Kv1.3通道控制的。我们有兴趣研究Kv1.3通道在T细胞和抗原提呈细胞接触点的位置如何影响通道功能的细节。为此，我们提出了一种结合纳米技术和人工抗原呈递细胞状表面的新方法。

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.