Single Molecule Imaging to Quantify FC??RI Signaling Dynamics

单分子成像量化 FC??RI 信号动态

• 批准号: 8627186
• 负责人: Diane Lidke
• 金额: $ 28.53万
• 依托单位: UNIVERSITY OF NEW MEXICO HEALTH SCIS CTR
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-01 至 2017-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8627186
• 关键词: Adaptor Signaling Protein; Affinity; Antigens; Arteriosclerosis; Asthma; Behavior; Binding; Biochemical; Biochemistry; Biological; Biology; Cancer Science; Cell Shape; Cells; Cellular biology; Color; Communication; Communities; Complex; Data; Development; Drug Design; Environment; Event; Fostering; Goals; Hypersensitivity; IgE Receptors; Image; Imaging Techniques; Immune response; Immune system; Kinetics; Knowledge; Ligand Binding; Lipid Bilayers; Malignant Neoplasms; Measures; Membrane; Methodology; Methods; Mission; Molecular; Outcome; Phosphorylation; Physiological; Process; Protein Binding; Protein Dynamics; Proteins; Public Health; Research; Residencies; Rest; Rheumatoid Arthritis; Role; Sentinel; Shapes; Signal Transduction; System; Techniques; Time; Work; base; crosslink; imaging modality; innovation; insight; mast cell; new technology; novel; palmitoylation; protein protein interaction; receptor; receptor function; response; scaffold; segregation; single molecule

DESCRIPTION (provided by applicant): A key player in mast cell activation is the high affinity IgE receptor, Fc5RI. While much is known about the chain of events initiated by crosslinking of Fc?RI through multivalent antigen, there is a fundamental gap in our understanding of how protein dynamics facilitate signaling. Our long term goal is to understand how the dynamic and stochastic behavior of protein-protein interactions influences signal propagation. The objective of this proposal is to quantify dynamic protein interactions during early events in Fc?RI signaling. Our central hypothesis is that the duration of protein-protein interactions modulates the signaling outcome. The rationale for the proposed research is that understanding the role of protein-protein interaction dynamics in signaling is the next step in understanding how the cell shapes the strength and quality of an immune response. We will integrate multiple imaging modalities, both novel and established, to capture and quantify early events that govern Fc?RI membrane associated signaling in order to achieve three specific aims: 1) To determine the interaction lifetime and diffusional dynamics that facilitate Fc?RI activation; 2) To quantify the kinetics of Syk adaptor protein binding to the signaling complex; and 3) To determine the cluster stability of LAT and the interplay between Fc?RI and LAT signaling patches. The approach is innovative because we will develop methodology for imaging biochemical events at the molecular level that will allow us to obtain dynamic information about signaling events that cannot be determined using traditional biochemistry techniques. The proposed research is significant because the quantitative information that we propose to obtain has not been directly measured before and will bring new perspectives to the cell biology community. Furthermore, the development of multi-color single molecule imaging techniques described here will be applicable to many other fundamental biological questions. The information we gain about Fc5RI signaling will help to fill the gaps in our knowledge of how Fc5RI initiates signaling. Ultimately, we expect that this information will open new avenues for drug design that target protein interactions and localization.

描述（由申请人提供）：肥大细胞激活的关键因素是高亲和力 IgE 受体 Fc5RI。虽然我们对 Fc?RI 通过多价抗原交联引发的一系列事件了解很多，但我们对蛋白质动力学如何促进信号传导的理解存在根本差距。我们的长期目标是了解蛋白质-蛋白质相互作用的动态和随机行为如何影响信号传播。该提案的目的是量化 Fc?RI 信号传导早期事件期间的动态蛋白质相互作用。我们的中心假设是蛋白质-蛋白质相互作用的持续时间调节信号传导结果。这项研究的基本原理是，了解蛋白质-蛋白质相互作用动力学在信号传导中的作用是了解细胞如何塑造免疫反应的强度和质量的下一步。我们将整合多种新颖和成熟的成像方式来捕获和量化控制 Fc?RI 膜相关信号传导的早期事件，以实现三个具体目标：1) 确定促进 Fc?RI 激活的相互作用寿命和扩散动力学； 2) 量化Syk接头蛋白与信号复合物结合的动力学； 3)确定LAT的簇稳定性以及Fc?RI和LAT信号转导补丁之间的相互作用。该方法具有创新性，因为我们将开发在分子水平上对生化事件进行成像的方法，这将使我们能够获得有关使用传统生化技术无法确定的信号事件的动态信息。拟议的研究意义重大，因为我们提出获得的定量信息以前从未被直接测量过，并将为细胞生物学界带来新的视角。此外，这里描述的多色单分子成像技术的发展将适用于许多其他基本生物学问题。我们获得的有关 Fc5RI 信号传导的信息将有助于填补我们关于 Fc5RI 如何启动信号传导的知识空白。最终，我们预计这些信息将为针对蛋白质相互作用和定位的药物设计开辟新途径。