EGF RECEPTOR ACTIVATION AND INTERACTION WITH ERBB FAMILY RECEPTORS

EGF 受体激活以及与 ERBB 家族受体的相互作用

• 批准号: 8065915
• 负责人: Linda Joy Pike
• 金额: $ 27.56万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-18 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8065915
• 关键词: Adopted; Affinity; Agonist; Behavior; Binding; Binding Proteins; Binding Sites; Cell Signaling Process; Cells; Characteristics; Clinical; Complex; Data; Development; Dissociation; EGF gene; Enzymes; Epidermal Growth Factor Receptor; Equilibrium; ErbB Receptor Family Protein; ErbB4 gene; Family; Family member; Fluorescence; Foundations; Goals; Heterodimerization; Heterogeneity; Homo; Human; Investigation; Lateral; Lead; Life; Ligand Binding; Ligands; Measures; Mediating; Methods; Modeling; Mutate; Nature; Pathway interactions; Pattern; Process; Proteins; Receptor Activation; Receptor Protein-Tyrosine Kinases; Role; Signal Pathway; Signal Transduction; Site; Spectrum Analysis; Stream; System; Treatment Efficacy; dimer; discount; expectation; insight; member; monomer; novel; receptor; receptor binding; therapeutic target; tumor

The EGF receptor is the first member of the ErbB family of growth factor receptor tyrosine kinases that also includes ErbB2, ErbB3 and ErbB4. Binding of EGF to its receptor induces the formation of EGF receptor homodimers, resulting in autophosphorylation of the EGF receptor. The EGF receptor is able to heterodimerize with other members of the ErbB family, with ErbB2 being the preferred heterodimerization partner. Recent data suggest that ErbB receptors may form heterotetramers and that activated oligomers may dissociate into monomers that re-dimerize with inactive monomers in a process referred to as secondary dimer formation. EGF shows heterogeneity in affinity when binding to its receptor. This observation has been attributed to the existence of two independent classes of sites. How these arise from the expression of a single EGF receptor protein has never been explained. Recently, we have shown that EGF binding can be described by a model that involves negative cooperativity in an aggregating system. In this model, the affinity of EGF for a monomer and an unoccupied dimer is similar. However, binding to the second site on an EGF receptor dimer is negatively cooperative. This model predicts that the negative cooperativity will lead to EGF-induced dissociation of the dimer at high concentrations of EGF, a prediction that we have confirmed using fluorescence correlation spectroscopy and enzyme complementation to examine EGF receptor oligomerization. Our long-term goal is to understand how ErbB receptors are activated and to develop a model for predicting the interactions among ErbB family members. Using our new model as the starting point for further investigations, the specific aims of this proposal are to: 1) Quantify the interaction of the EGF receptor and ErbB2 through ligand binding studies; 2) Determine whether EGF receptor-mediated activation of ErbB2 occurs via the formation of a hetero-tetrameric complex and/or involves dissociation of dimeric receptor complexes; and, 3) Determine whether different EGF receptor ligands induce characteristic patterns of EGF receptor oligomers and whether this can be related to differences in downstream signaling. EGF equilibrium binding studies will be used to quantitatively characterize the interaction of the EGF receptor with ErbB2. Fluorescence correlation spectroscopy and brightness analysis along with enzyme complementation will be employed to assess the oligomerization state of EGFR/ErbB2 heteromeric complexes while chemically-induced dimer formation will be used to probe the activation mechanism of ErbB2 within these complexes. A combination of these methods will be applied to assess oligomerization of the EGF receptor elicited by three different agonists. Signaling stimulated by the three agonists will be assessed by measuring the activation of downstream pathways. Together, these data will characterize the interactions of the EGF receptor and ErbB2 allowing the development of a quantitative model for the activation of ErbB2 via the EGF receptor and provide insight into the role of ErbB family oligomers in the process of cell signaling.

EGF受体是生长因子受体酪氨酸激酶ErbB家族中的第一个成员， 还包括ErbB2、ErbB3和ErbB4。EGF与其受体的结合诱导EGF受体的形成 同源二聚体，导致EGF受体的自动磷酸化。EGF受体能够异源二聚体 与ErbB家族的其他成员，ErbB2是首选的异二聚化伙伴。近期 数据表明，ErbB受体可能形成异四聚体，激活的寡聚体可能解离成 在称为二次二聚体形成的过程中与不活跃的单体重新聚合的单体。 当EGF与其受体结合时，其亲和力表现出异质性。这一观察结果被归因于 存在两类独立的站点。这些是如何从单个EGF的表达中产生的 受体蛋白从未被解释过。最近，我们已经证明EGF结合可以用以下公式来描述 一种在聚合系统中涉及负协作性的模型。在这个模型中，EGF对一种 单体和未被占据的二聚体相似。然而，与EGF受体二聚体上的第二个位点结合 是消极合作的。该模型预测，负的协同性将导致EGF诱导的解离 在高浓度的EGF下的二聚体，我们已经用荧光证实了这一预测 相关光谱和酶互补法检测EGF受体寡聚。 我们的长期目标是了解ErbB受体是如何激活的，并开发一种模型 预测ErbB家族成员之间的相互作用。使用我们的新模式作为进一步发展的起点 研究表明，这一建议的具体目的是：1)量化EGF受体和 2)确定EGF受体是否介导了ErbB2的激活 通过形成异四聚体复合体和/或涉及二聚体受体的解离而发生 复合体；以及，3)确定不同的EGF受体配体是否诱导EGF的特征模式 受体寡聚体以及这是否与下游信号的差异有关。 EGF平衡结合研究将用于定量描述EGF的相互作用 受体与ErbB2结合。荧光相关光谱和亮度分析与酶 互补作用将被用来评估EGFR/ErbB_2杂化络合物的齐聚状态 而化学诱导的二聚体的形成将被用来探索ErbB2在这些细胞中的激活机制 复合体。这些方法的组合将被应用于评估EGF受体的寡聚化 由三种不同的激动剂引起。由三种激动剂刺激的信号将通过测量 下游通路的激活。总之，这些数据将表征EGF的相互作用 受体和ErbB2可通过EGF建立ErbB2激活的定量模型 并深入了解ErbB家族寡聚体在细胞信号转导过程中的作用。