Proteasome Regulation of Interleukin-5 Receptor Endocyt*

描述（由申请人提供）：白细胞介素-5 （IL-5）是一种造血细胞因子，在特应性和哮喘中嗜酸性粒细胞炎症的病理生理中起核心作用。了解消除IL-5受体（IL-5R）诱导的炎症信号的机制对于开发调节IL-5介导的炎症的新方法具有重要意义。本应用程序的目的是确定通过泛素和蛋白酶体降解途径控制IL-5R内化的分子机制。我们的总体假设是，il -5诱导的IL-5R内化受到两种不同机制的调节：泛素/蛋白酶体降解途径和网格蛋白介导的内吞作用。目的1。验证蛋白酶体通过首先降解(c)细胞质结构域介导连接的IL-5R内化的假设。为了确定部分(c)胞质结构域的降解是否是IL-5R内化的起始信号，将检查各种细胞质截断的(c)突变体介导连接的IL-5R的蛋白酶体非依赖性内化的能力。此外，还将研究其他参与c的细胞因子IL-3和GM-CSF对c内化过程中蛋白酶体活性的要求。目标2。验证IL-5R通过网格蛋白介导的内吞作用内化的假设，并且受体的内化发生在蛋白酶体降解(c)细胞质结构域之前。将使用两种不同的方法来抑制网格蛋白介导的内吞作用，以确定IL-5R是否通过这种机制内化，以及il -5刺激的c信号传导和蛋白酶体降解是否在IL-5R内化后发生。目标3。探讨c泛素化在IL-5R内化中的功能作用和性质。温度敏感的ts20细胞系在泛素结合上有温度敏感缺陷，(c突变体在所有形式的泛素化上都有缺陷，以及显性的阴性泛素异构体将被用来研究(c泛素化在IL-5R内吞作用中的作用和性质。

DESCRIPTION (provided by applicant): Interleukin-5 (IL-5) is a hematopoietic cytokine that is central to the pathophysiology of eosinophilic inflammation in atopy and asthma. Understanding mechanisms that extinguish IL-5 receptor (IL-5R)-induced inflammatory signals has importance for the development of novel approaches to modulate IL-5-mediated inflammation. The goal of this application is to define the molecular mechanisms controlling IL-5R internalization by the ubiquitin and proteasome degradation pathway. Our overall hypothesis is that IL-5-induced IL-5R internalization is regulated by two different mechanisms: the ubiquitin/proteasome degradation pathway and clathrin-mediated endocytosis. Aim 1. Test the hypothesis that proteasomes mediate internalization of the ligated IL-5R by first degrading the (c cytoplasmic domain. To determine if degradation of a portion of the (c cytoplasmic domain is the initiating signal for IL-5R internalization, various cytoplasmically truncated (c mutants will be examined for their ability to mediate proteasome-independent internalization of the ligated IL-5R. In addition, requirement for proteasome activity in (c internalization by the other (c engaging cytokines, IL-3 and GM-CSF, will be investigated. Aim 2. Test the hypothesis that the IL-5R is internalized by clathrin-mediated endocytosis and that internalization of the receptor occurs before proteasome degradation of the (c cytoplasmic domain. Two different approaches that inhibit clathrin-mediated endocytosis will be used to determine if the IL-5R is internalized by this mechanism and if IL-5-stimulated (c signaling and proteasome degradation occur after IL-5R internalization. Aim 3. Investigate the functional role and nature of (c ubiquitination in IL-5R internalization. The temperature sensitive ts20 cell line which has a temperature sensitive defect in ubiquitin conjugation, a (c mutant that is defective in all forms of ubiquitination, and dominant negative ubiquitin isoforms will be used to examine the role and nature of (c ubiquitination in IL-5R endocytosis.