Control of ErbB Receptor Sorting in Endosomes

内体中 ErbB 受体分选的控制

• 批准号: 6794606
• 负责人: CATHLEEN R CARLIN
• 金额: $ 27.54万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-05 至 2006-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6794606
• 关键词: biological signal transduction; cell line; cell membrane; electron microscopy; gene mutation; intracellular transport; ligands; lysosomes; membrane fusion; membrane proteins; mitogen activated protein kinase; nuclear magnetic resonance spectroscopy; phosphorylation; phosphotransferases; protein protein interaction; protein structure function; receptor binding; receptor expression; receptor mediated endocytosis; site directed mutagenesis; vesicle /vacuole

DESCRIPTION (provided by applicant): The endosomal apparatus is a major site of membrane sorting in both endocytic and exocytic pathways. For ligand activated receptors it also provides a mechanism for achieving the proper balance of cell signaling from different membrane compartments. A main objective of this proposal is to understand how endocytosis controls cellular responses to the EGF receptor (EGFR), the prototype for the ErbB receptor family. The complexity of endocytic trafficking predicts that EGFRs use a multitude of distinct sorting signals at different locations in the cell. Characterization of these signals provides a framework for understanding regulation of EGFR transit in the endosomal apparatus. We have identified three non-overlapping sorting signals that control critical benchmarks in EGFR trafficking: 679-LL, which controls sorting in multivesicular endosome-to-lysosome transport intermediates or MVEs; 954-YLVI, which controls lysosomal sorting at a site intersecting the exocytic pathway; and 662-RxxxxPLTP, which controls sorting from endosomes to the plasma membrane. Completion of the proposed studies will provide new insights on the coordinated action of these signals during ligand-regulated EGFR trafficking. Since related ErbB receptors have divergent sorting signals compared to EGFR, we will also gain insight to how ErbB signaling in general is controlled by intracellular trafficking. This is particularly important for ErbB2, whose ligand-dependent activation by EGFR requires EGFR sequences that regulate post-endocytic trafficking and not intrinsic tyrosine kinase activity. The availability of dominant-inhibitory mutations that disrupt EGFR trafficking at defined steps also provides a unique set of reagents with which to study the temporal and spatial organization of signaling from endosomes. The following hypotheses will be tested: 1. Ligand-induced EGFR post-endocytic sorting to lysosomes is mediated by the concerted action of multiple sorting signals acting in a step-wise fashion. 2. The 679-LL MVE sorting signal is recognized as part of a larger motif whose physiological function is dependent on protein-protein interactions. 3. The 662-RxxxxPLTP sorting signal controls EGFR recycling to the plasma membrane, and is regulated by MAP kinase-mediated phosphorylation of Thr669. 4. The 679-LL sorting signal regulates EGFR signaling by controlling the balance of growth stimulatory pathways elicited during endocytosis. This hypothesis is based on new data suggesting that EGFRs with an inactivating L679A,L680A mutation selectively activate survival pathways compared to wild-type, and will be tested using both in vitro and in vivo experimental models.

描述(由申请人提供)：内膜器是胞内和胞外途径中膜分离的主要部位。对于配体激活的受体，它还提供了一种机制，以实现来自不同膜间隔的细胞信号的适当平衡。这项建议的一个主要目的是了解内吞作用如何控制细胞对ErbB受体家族的原型--EGF受体(EGFR)的反应。内吞转运的复杂性表明，EGFR在细胞内的不同位置使用大量不同的分选信号。这些信号的特征为理解EGFR在内体器官中转运的调控提供了一个框架。我们已经确定了三个不重叠的分选信号，它们控制着EGFR转运中的关键基准：679-LL，它控制着多囊内小体到溶酶体的转运中间体或VME的分选；954-YLVI，它控制着溶酶体在胞外途径的交叉处的分选；以及662-RxxxxPLTP，它控制着从内小体到质膜的分选。拟议研究的完成将为这些信号在配体调节的EGFR贩运过程中的协调行动提供新的见解。由于相关的ErbB受体与EGFR相比具有不同的分选信号，我们还将深入了解ErbB信号一般是如何由细胞内转运控制的。这对ErbB2尤其重要，它的配体依赖于EGFR的激活需要EGFR序列来调节内吞后转运，而不是内在的酪氨酸激酶活性。显性抑制突变的可用性在确定的步骤上扰乱了EGFR的运输，也提供了一套独特的试剂来研究来自内体的信号的时间和空间组织。将检验以下假设：1.配体诱导的EGFR对溶酶体的内吞后分选是由多个分选信号以分步方式协同作用而介导的。2.679-L1 MVE分选信号被认为是一个更大的基序的一部分，其生理功能依赖于蛋白质-蛋白质的相互作用。3.662-RxxxxPLTP分选信号控制EGFR向质膜的循环，并受MAP激酶介导的Thr669磷酸化的调节。4.679-L1分选信号通过控制内吞过程中的生长刺激通路的平衡来调节EGFR信号转导。这一假说基于新的数据，该数据表明，与野生型相比，具有失活L679A和L680A突变的EGFR选择性地激活生存途径，并将使用体外和体内实验模型进行测试。