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Red and Far-Red Fluorescent Biosensors for Compartmentalized Redox Signaling

用于分隔氧化还原信号传导的红色和远红荧光生物传感器

基本信息

批准号：
10222722
负责人：
Huiwang Ai
金额：
$ 37.84万
依托单位：
UNIVERSITY OF VIRGINIA
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-05 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10222722
关键词：
Aging Animal Model Basic Science Biology Biosensor Cancer Etiology Cell Compartmentation Cell Nucleus Cell model Cells Chemicals Color Coupled Coupling Cytosol Detection Development Disease Doxorubicin Drug Screening Endoplasmic Reticulum Engineering Epidermal Growth Factor FDA approved Fluorescence Glutathione Goals Human Hydrogen Peroxide Image Imaging Device Individual Inflammatory Intervention Laboratories Lead Libraries Light Measurement Measures Methods Mitochondria Molecular Monitor Nitrogen Nuclear Oncology Organism Oxidation-Reduction Oxidative Stress Oxygen Penetration Pharmaceutical Preparations Phosphotransferases Photons Physiological Process Protein Disulfide Isomerase Proteins Regulation Reporting Research Sensory Signal Transduction Stimulus Sulfhydryl Compounds Sulfur TXN gene Technology Tissues Toxicology Work absorption animal imaging base biomaterial compatibility chemotherapy multiplexed imaging mutant novel novel therapeutics oxidation public health relevance ratiometric red fluorescent protein response sensor spatiotemporal subcellular targeting technology research and development tool toxicant translational study whole animal imaging

项目摘要

ABSTRACT Redox biology has been increasingly recognized as one of the key themes in cell signaling. A major obstacle in this field is the lack of capabilities to monitor and measure clearly defined redox processes in live cells (including subcellular domains in live cells), tissues, or organisms. The goal of this 4-year project, submitted in response to PAR-17-045 “Focused Technology Research and Development”, is to expand the fluorescent protein toolbox for red and far-red fluorescent biosensors that can be used to image compartmentalized, thiol-based redox signaling in live cells and organisms. In this project, we aim to advance the fluorescent biosensor technology in the following aspects : (1) we will expand fluorescent-protein-based biosensors for additional redox parameters, including the redox of thioredoxin and protein disulfide isomerase (PDI) in various cell compartments, such as the nucleus, cytosol, mitochondria, and endoplasmic reticulum (ER); (2) we will expand the colors of fluorescent-protein-based redox biosensors to red and far-red; (3) we will validate these new biosensors and demonstrate the use of fluorescent-protein-based biosensors in compatible colors for multicompartment/multiplex imaging. Because UV or blue light, which can induce intracellular oxidation, is not needed to excite red and far- red fluorescent biosensors, our work will increase the biocompatibility of fluorescent-protein-based redox biosensors. Furthermore, red and far-red biosensors will assist in whole-animal imaging, since compared to green ones, they reduce tissue absorption and scattering. This project will lead to important imaging tools that may accelerate the understanding of redox regulation and signaling. As with the transformative impact of fluorescent-protein-based kinase sensors, we expect that these redox biosensors will enable and transform a large array of basic and translational studies.

摘要氧化还原生物学已越来越被认为是细胞信号转导的关键主题之一。一位少校这一领域的障碍是缺乏实时监控和测量明确定义的氧化还原过程的能力细胞(包括活细胞中的亚细胞域)、组织或生物体。这个为期4年的项目的目标是，为响应PAR-17-045《重点技术研究与开发》而提交，旨在扩展可用于成像的红色和远红荧光生物传感器的荧光蛋白工具箱活细胞和生物体中基于硫醇的分段氧化还原信号。在本项目中，我们的目标是在以下几个方面推进荧光生物传感器技术：(1)我们将扩展基于荧光蛋白的生物传感器，以获得更多的氧化还原参数，包括硫氧还蛋白和蛋白质二硫键异构酶(PDI)在不同的细胞室，如细胞核，胞浆，线粒体、内质网(ER)；(2)我们将扩展以荧光蛋白为基础的颜色将生物传感器氧化还原为红色和远红色；(3)我们将验证这些新的生物传感器并演示基于荧光蛋白的生物传感器，具有兼容的颜色，可用于多隔室/多路成像。因为可以诱导细胞内氧化的紫外线或蓝光不需要激发红光和远红光。红色荧光生物传感器，我们的工作将提高基于荧光蛋白的氧化还原的生物相容性生物传感器。此外，红色和远红色生物传感器将有助于整个动物的成像，因为与绿色的，它们可以减少组织的吸收和散射。这个项目将导致重要的成像工具，可能会加速对氧化还原的理解监管和信号传递。与基于荧光蛋白的激酶传感器的变革性影响一样，我们预计这些氧化还原生物传感器将使大量的基础和转化性研究得以实现和转变。