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Illuminating Dynamic Receptor Clustering in the Epidermal Growth Factor Receptor

照亮表皮生长因子受体中的动态受体簇

基本信息

批准号：
8444530
负责人：
Bjoern Markus Reinhard
金额：
$ 30.74万
依托单位：
BOSTON UNIVERSITY (CHARLES RIVER CAMPUS)
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-06-01 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8444530
关键词：
Affinity Behavior Binding Blinking Cancer Diagnostics Cancerous Cell Line Cell Surface Receptors Cell membrane Cells Coupling Development EGF gene Energy Transfer Epidermal Growth Factor Epidermal Growth Factor Receptor Fluorescence Fluorescence Microscopy Fluorescence Resonance Energy Transfer Goals Gold Higher Order Chromatin Structure Imaging Device Imaging technology Individual Investigation Label Length Life Ligands Link Malignant Neoplasms Measurement Mediation Metals Microscopy Molecular Monitor Motion Nanotechnology Pathway interactions Play Process Public Health Receptor Activation Receptor Signaling Regulation Relative (related person)Resolution Role Side Signal Transduction Signal Transduction Pathway Spatial Distribution Speed Structure Surface System Technology Therapeutic Intervention Time anti-cancer therapeutic attenuation base cancer cell cancer therapy dimer fluorescence imaging improved molecular dynamics nanometer nanoparticle new technology overexpression particle receptor tool tumor

项目摘要

PROJECT SUMMARY/ABSTRACT Illuminating Dynamic Receptor Clustering in the Epidermal Growth Factor Receptor Signal Transduction Pathway Using Plasmon Coupling Aberrant Epidermal Growth Factor Receptor (EGFR) activity has been linked to tumor formation and progression in malignant cells. A molecular understanding of the EGFR signaling mechanism offers opportunities for the development of efficient anti-cancer therapeutic strategies. The EGFR activation and its mediation by interreceptor ectodomain interactions, however, still pose many questions. The operational form in the EGFR signal transduction pathway is not the individual EGF receptor, but dimers and potentially higher order oligomers or even larger functional units, referred to as clusters. The influence of transient interactions between EGFRs in clusters with dynamic short- and long-range orders could play a prominent role in the regulation of the EGFR pathway. To elucidate the connections between receptor cell surface organization, receptor dynamics, and the activation, progression, attenuation and therapeutic intervention of signaling distances between individual EGFRs on the 1-100 nm length scale need to be monitored with high temporal resolution. This distance range lies in the "resolution gap" of conventional fluorescent microscopy which is defined by the spatial Fluorescence Resonance Energy Transfer barrier of 10 nm on one side and the diffraction resolution limit in the visible of ~300 nm on the other side. Instead of using a fluorescence microscopy this proposal aims to unravel the dynamics of EGFR oligomerization and clustering using plasmon coupling between gold nanoparticle labeled EGFRs in living cells in real time. The advantages of plasmon coupling microscopy are given by the photophysical stability of the probes, noble metal nanoparticles don't blink or bleach, their signal intensity, and the fact that plasmons in close by particles couple. This plasmon coupling enables to detect distances and distance changes significantly beyond the spatial FRET barrier and enables distance measurements in the resolution gap of fluorescence microscopy. The specific aims of this project are: 1. Develop a new molecular ruler that allows monitoring distances between EGFRs on living cells in real time on length scales between 1 - 100 nm. This new technology will enable us to probe both the short- and long-range order of EGFRs on the surface of living cells. 2. Experimentally verify the hypothesis that EGFRs are organized in clusters with inter-EGFR distances ranging from nanometers to tens of nanometers. Monitor changes in the size and spatial distribution of EGFR clusters in living cells in real time upon addition of EGF. 3. Monitor inter-EGFR distances within the clusters and experimentally verify the hypothesis that EGF induces changes in the inter-EGFR distances.

项目总结/文摘