Secretase Processing of ErbB-4 Receptor Tyrosine Kinase

ErbB-4 受体酪氨酸激酶的分泌酶加工

• 批准号: 6506422
• 负责人: GRAHAM F CARPENTER
• 金额: $ 30.14万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2005-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6506422
• 关键词: bioassay; biological signal transduction; cell line; chemical cleavage; enzyme activity; enzyme inhibitors; enzyme mechanism; fluorescence resonance energy transfer; gene mutation; gene targeting; genetically modified animals; growth factor receptors; laboratory mouse; metalloendopeptidases; microarray technology; neoplastic cell; phosphorylation; presenilin; protein localization; protein structure function; protein transport; protein tyrosine kinase; proteolysis; receptor expression; transcription factor; yeast two hybrid system

DESCRIPTION (provided by applicant): The focus of this application is to explore a novel pathway for the proteolytic processing of receptor tyrosine kinases. In particular, this involves the two step cleavage of the ErbB-4 receptor that binds the growth factor heregulin. Cleavage of ErbB-4 can be stimulated by heregulin or TPA and results in the relocalization of the ErbB-4 cytoplasmic domain, including its tyrosine kinase, from the plasma membrane to the nucleus. The application seeks to elucidate the biochemical parameters of the two cleavage events, as well as the biological significance of the processing pathway for ErbB-4 mediated cellular responses. The first aim will characterize the initial and essential secretase (metalloprotease)-dependent cleavage of the ErbB-4 ectodomain. This will include identification of the cleavage site, construction of receptor and protease mutants, evaluation of the contributions of ErbB-4 structure to protease recognition, and the relationship of cleavage and signal transduction to heregulin-induced translocation of ErbB-4 to membrane microdomains. The second aim will concentrate on the subsequent gamma-secretase (presenilin) mediated cleavage that releases the ErbB-4 cytoplasmic domain from the plasma membrane. This will include analysis of the gamma-secretase cleavage site, the role of PDZ domain proteins and presenilins, and nuclear translocation of the ErbB-4 cytoplasmic domain. The final aim is constructed to address the physiological significance of ErbB-4 cleavage and nuclear re-localization of the cytoplasmic domain. This will include cell biological experiments of cytoplasmic domain functions in the nucleus, such as tyrosine phosphorylation and the activation of gene expression. Selective inhibitors of proteolytic processing will be used to assess the role of this pathway in ErbB-4 mediated biological responses. Lastly, a mouse "knockin" approach is proposed to assess the physiological significance of this novel receptor processing pathway in the animal.

描述（由申请人提供）：本申请的重点是探索受体酪氨酸激酶蛋白水解加工的新途径。特别是，这涉及ERBB-4受体的两个步骤裂解，该受体结合了这里的生长因子。 ERBB-4的裂解可以通过这里肿瘤或TPA刺激，并导致ERBB-4细胞质结构域的重新定位，包括其酪氨酸激酶，从质膜到核。该应用程序旨在阐明两个裂解事件的生化参数，以及ERBB-4介导的细胞反应的加工途径的生物学意义。 第一个目的将表征ERBB-4胞外域的初始和必需的泌尿酶（金属蛋白酶）依赖性裂解。这将包括识别裂解位点，受体和蛋白酶突变体的构建，评估ERBB-4结构对蛋白酶识别的贡献，以及裂解和信号转导与此处糖蛋白诱导的ERBB-4对膜微域的易位的关系。第二个目标将集中于随后的伽马分泌酶（Presenilin）介导的裂解，从而从质膜释放ERBB-4细胞质结构域。这将包括分析γ-分泌酶裂解位点，PDZ结构蛋白和presenilins的作用以及ERBB-4细胞质结构域的核转运。最终目标是解决ERBB-4裂解和细胞质结构域的核重新定位的生理意义。这将包括细胞胞质结构域功能的细胞生物学实验，例如酪氨酸磷酸化和基因表达的激活。蛋白水解处理的选择性抑制剂将用于评估该途径在ERBB-4介导的生物学反应中的作用。最后，提出了一种小鼠的“敲蛋白”方法来评估动物中这种新型受体加工途径的生理意义。