Collaborative Research: Modeling Spatiotemporal Control of EGFR-ERK Signaling in Gene-edited Cell Systems

合作研究：基因编辑细胞系统中 EGFR-ERK 信号传导的时空控制建模

• 批准号: 1715132
• 负责人: Alexander Sorkin
• 金额: $ 53.13万
• 依托单位: University of Pittsburgh
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1715132&HistoricalAwards=false
• 关键词: Collaborative; Research; Modeling; Spatiotemporal; Control

In this project, the investigators will develop a new experimental and computational platform to answer the longstanding biological question of how cellular receptors generate biochemical signals as they move among various subcellular compartments. By helping to quantitatively resolve certain apparently conflicting observations that have recently been made within a biological system previously thought to be well understood, this project will add fundamentally new understanding of basic mechanisms of cell signaling that may have implications across numerous receptor systems in cell biology. Moreover, due to the broadly important role that receptors, and the biochemical signals they generate, play in virtually every biological process, the new quantitative understanding gained will ultimately impact ongoing efforts to optimally engineer receptor-mediated signaling. This project also includes a broad and detailed set of integrated educational objectives that leverage the specific scientific objectives and activities of this project to reach students at different training stages and from diverse backgrounds. These education and outreach activities involve innovation upon existing programs at the partnering institutions as well as the creation of new outreach efforts that will target larger audiences, such as a summer academy of science for high school students and a collaboration with the Museum of Science and Industry in Tampa, FL to develop a live interaction program on cell signaling. The biochemical signaling that results upon ligand binding to receptor tyrosine kinases is accompanied by receptor clustering and internalization, or endocytosis. While it has been understood for decades that endocytosis enables cells to degrade or recycle receptors, the question of how endocytosis ultimately impacts downstream signal propagation remains wide open. For example, even for the highly studied epidermal growth factor receptor (EGFR) the literature contains highly cited, yet conflicting reports that conclude that EGFR endocytosis either promotes or impedes downstream signaling through the extracellular regulated kinase (ERK) pathway. In this project, the investigators will explore the hypothesis that the intrinsic complexity of the spatiotemporal signaling regulatory mechanisms creates parameter spaces for both answers to be true depending on the cellular context. The complexity of the receptor trafficking and signaling system cannot be fully dissected, however, without integrating quantitative experimental measurements that faithfully reflect cell biology with spatiotemporal computational models. Accordingly, the overarching goal of this project is to combine novel live-cell imaging of gene-edited cell systems with mechanistic models that capture system complexity with unprecedented detail to resolve the kinds of conflicting reports previously mentioned, as well as more recent surprising observations that certain proteins conventionally thought to remain in complex during signal transduction may become physically separated through trafficking processes. Ultimately, the proposed integration of gene-edited cell systems, quantitative live-cell imaging and biochemical measurements, and a spatiotemporal computational model will lead to an entirely new quantitative platform to answer these kinds of questions for EGFR and other receptor signaling systems.

在这个项目中，研究人员将开发一个新的实验和计算平台，以回答长期存在的生物学问题，即细胞受体如何在不同的亚细胞间移动时产生生化信号。通过帮助定量地解决最近在以前被认为很好地理解的生物系统内所做的某些明显相互矛盾的观察，该项目将从根本上增加对细胞信号的基本机制的理解，这些机制可能会对细胞生物学中的许多受体系统产生影响。此外，由于受体及其产生的生化信号在几乎每个生物过程中都扮演着广泛重要的角色，新的量化理解最终将影响正在进行的优化设计受体介导的信号传递的努力。该项目还包括一套广泛而详细的综合教育目标，利用该项目的具体科学目标和活动来接触处于不同培训阶段和不同背景的学生。这些教育和外展活动包括对合作机构现有项目的创新，以及创建针对更多受众的新外展努力，例如为高中生设立夏季科学学院，以及与佛罗里达州坦帕市的科学与工业博物馆合作开发关于细胞信号的现场互动项目。配体与受体酪氨酸激酶结合所产生的生化信号伴随着受体聚集和内化，或内吞作用。虽然几十年来人们一直了解内吞作用使细胞能够降解或回收受体，但内吞作用最终如何影响下游信号传播的问题仍然悬而未决。例如，即使对于高度研究的表皮生长因子受体(EGFR)，文献中也包含大量被引用但相互矛盾的报告，结论是EGFR内吞作用通过细胞外调节激酶(ERK)途径促进或阻碍下游信号传递。在这个项目中，研究人员将探索这样一个假设，即时空信号调节机制的内在复杂性根据细胞背景为两个答案都为真创造了参数空间。然而，如果不将真实反映细胞生物学的定量实验测量与时空计算模型结合起来，就不能完全剖析受体运输和信号系统的复杂性。因此，该项目的首要目标是将基因编辑细胞系统的新型活细胞成像与机械模型相结合，这些模型以前所未有的细节捕捉系统的复杂性，以解决前面提到的各种相互矛盾的报告，以及最近令人惊讶的观察结果，即某些传统上认为在信号转导过程中保持复杂状态的蛋白质可能会通过贩运过程在物理上分离。最终，基因编辑细胞系统、定量活细胞成像和生化测量以及时空计算模型的提出将为EGFR和其他受体信号系统带来一个全新的定量平台来回答这些问题。

项目成果

Localization dynamics of endogenous fluorescently labeled RAF1 in EGF-stimulated cells

EGF 刺激细胞中内源性荧光标记 RAF1 的定位动态

• DOI: 10.1091/mbc.e18-08-0512
• 发表时间: 2019
• 期刊: Molecular Biology of the Cell
• 影响因子: 3.3
• 作者: Surve, Sachin V.;Myers, Paul J.;Clayton, Samantha A.;Watkins, Simon C.;Lazzara, Matthew J.;Sorkin, Alexander;Heldin, Carl-Henrik
• 通讯作者: Heldin, Carl-Henrik