Regulation of EGF receptor trafficking fate by phosphorylation and Odin

生长因子与受体的结合是调节后生动物生长发育的关键机制。表皮生长因子受体(EGFR)是典型的生长因子受体。活化的EGFR通过内吞作用迅速内化。内化的EGFR要么循环回到细胞表面，要么在溶酶体中降解，这是运输分选开关的结果。EGFR也内化和循环，以响应细胞压力。这些过程控制细胞表面的受体密度，进而对不同的配基浓度梯度和环境信号做出反应，这些在后生动物中是高度可变的。我们的长期目标是从定量机制的角度了解动态共价修饰和蛋白质相互作用如何控制生长因子受体的运输命运。 我们发现EGFR磷酸化(Y998，S1039)和Odin蛋白是影响EGFR转运的因素。例如，在Y998未能将激活的EGFR磷酸化会导致回收，而没有ODIN的细胞则缺乏EGFR回收。受体转运的调控已被深入研究，但EGFR磷酸化如何调控其循环或降解转运命运的分子机制细节尚不清楚。我们的近期目标是确定构成循环或降解开关的磷酸化介导的EGFR分类的分子机制。 为了实现这一目标，我们提出了以下具体目标： 目标1：解决测试Odin在EGFR回收中的作用的问题：(A)Odin是否结合Y998进行回收，但不结合刺激降解的PY998？(B)ODIN在应激诱导的EGFR循环中起作用吗？(C)ODIN的磷酸化是否调节其与EGFR的相互作用？ 目的：检测EGFR-S1039的磷酸化是否控制着EGFR的循环，并阻止其转运到溶酶体。(A)回收EGFR是否需要S1039？(B)受体相关的磷酸酶是否调节S1039的磷酸化，从而调节EGFR的循环？(C)确定EGFR-S1039磷酸化的时间和空间调节，以确定其在循环中的作用。 从机制上理解磷酸化/ODIN介导的EGFR分类将导致未来研究受体如何在复杂生物体中调节生长因子和环境信号。该计划还将有助于HQP在分子和细胞生物学以及蛋白质组学领域的培训。

Binding of growth factors to receptors is a key mechanism that regulates the growth and development of metazoans. Epidermal growth factor receptor (EGFR) is a prototypical growth factor receptor. Activated EGFR is rapidly internalization by endocytosis. Internalized EGFR either recycle back to the cell surface or degrade in lysosomes as a consequence of a trafficking sorting switch. EGFR also internalize and recycle in response to cellular stresses. These processes control receptor density at the cell surface, and in turn signaling responses to diverse ligand concentration gradients and environmental cues, which are highly variable in metazoans.Our long term objective is to understand in quantitative mechanistic terms, how dynamic covalent modifications and protein interactions control the trafficking fate of growth factor receptors. We discovered EGFR phosphorylations (Y998, S1039) and the protein Odin as factors affecting EGFR trafficking. For example, failure to phosphorylate activated EGFR at Y998 causes recycling, whereas cells without Odin lack EGFR recycling. Regulation of receptor trafficking has been intensively studied, but molecular mechanistic details of how EGFR phosphorylation regulates its recycle-or-degrade trafficking fate are not known. Our near term objective is to determine the molecular mechanism of phosphorylation-mediated EGFR sorting that constitutes a recycle-or-degrade switch. To achieve this, we propose the following specific aims: Aim 1: Address questions that test Odin’s role in EGFR recycling: (A) Does Odin bind Y998 for recycling, but not pY998, which stimulates degradation? (B) Does Odin function in stress-induced EGFR recycling? (C) Does phosphorylation of Odin modulate its interaction with EGFR? Aim2: Test if EGFR-S1039 phosphorylation controls EGFR recycling, and prevents transport to the lysosome. (A) Is S1039 required for EGFR recycling? (B) Does a receptor-associated phosphatase modulate S1039 phosphorylation, and consequently EGFR recycling? (C) Determination of temporal and spatial regulation of EGFR-S1039 phosphorylation towards defining its role in recycling. A mechanistic understanding phosphorylation/Odin-mediated EGFR sorting will lead to future studies of how receptor trafficking modulates growth factor and environmental signals in complex organisms. This program will also contribute to the training of HQP in the fields of molecular and cell biology, and proteomics.