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

• 批准号: 9975915
• 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
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-05 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9975915
• 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

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 氧化还原生物学已经越来越被认为是细胞信号传导的关键主题之一。一个主要 该领域中的障碍是缺乏监测和测量活体中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结构域）、组织或生物体。这个为期四年的项目的目标， 提交的响应PAR-17-045“重点技术研究和开发”，是为了扩大 用于红色和远红色荧光生物传感器的荧光蛋白工具箱，可用于成像 在活细胞和生物体中的区室化的、基于硫醇的氧化还原信号传导。 在本项目中，我们的目标是在以下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）在各种细胞区室，如细胞核，胞质溶胶， 线粒体、内质网（ER）;（2）我们将扩大荧光蛋白的颜色基础 氧化还原生物传感器的红色和远红;（3）我们将验证这些新的生物传感器，并证明使用 基于荧光蛋白的生物传感器，颜色兼容，用于多室/多重成像。 因为可以诱导细胞内氧化的紫外光或蓝光不需要激发红光和远红光， 红色荧光生物传感器，我们的工作将增加生物相容性的荧光蛋白基氧化还原 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成像，因为与 绿色的，它们减少组织吸收和散射。 这个项目将导致重要的成像工具，可能会加速氧化还原的理解 调节和信号。与基于荧光蛋白的激酶传感器的变革性影响一样， 预计这些氧化还原生物传感器将使大量的基础和转化研究成为可能，并改变这些研究。