G-protein Coupled Receptor Mediated Directional Sensing

G蛋白偶联受体介导的定向传感

• 批准号: 6669979
• 负责人: Tian Jin
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6669979
• 关键词: Dictyostelium; G protein; G protein coupled receptor kinase; biological signal transduction; cell migration; cell motility; chemoattractants; chemotaxis; confocal scanning microscopy; cytokine receptors; fluorescence microscopy; fluorescence resonance energy transfer; intermolecular interaction; protein signal sequence; single cell analysis; tissue /cell culture

How do eukaryotic cells sense and move along a concentration gradient of chemical attractants? Chemoattractants such as chemokines are detected by G-protein coupled receptors (GPCR). Using a combination of molecular genetic, biochemical and cell biological approaches, we study chemoattractant elicited signal transduction in both mammalian cells and the model system Dictyostelium discoideum. We employ live cell imaging technology to visualize signaling events in single cells. During the last fiscal year, we established two fluorescence microscope systems: an Olympus IX70 equipped with two filter wheels that is linked to a coo-CCD digital camera and a Zeiss confocal Laser Scanning Microscope LSM 510 META. Using these systems, we are able to monitor the distribution of signaling proteins, tagged with various fluorescence proteins, in single living cells. In the mammalian system we determined the spatial distribution and visualized the signaling cascades of the chemokine receptor CXCR1 at the single cell level. In the D. discoideum system we monitored the temporal and spatial activities of GPCRs in the membranes of chemotaxing cells. Interactions between the subunits of the G proteins were detected as was GPCR-induced dissociation of G proteins in single cells.

真核细胞如何感知化学引诱剂的浓度梯度并沿着浓度梯度移动？ G 蛋白偶联受体 (GPCR) 可以检测趋化因子等趋化剂。我们结合分子遗传学、生物化学和细胞生物学方法，研究了哺乳动物细胞和盘基网柄菌模型系统中化学引诱剂引起的信号转导。我们采用活细胞成像技术来可视化单细胞中的信号事件。在上一财年，我们建立了两个荧光显微镜系统：一个配备两个滤光轮并连接到 coo-CCD 数码相机的 Olympus IX70 和一个蔡司共焦激光扫描显微镜 LSM 510 META。使用这些系统，我们能够监测单个活细胞中标记有各种荧光蛋白的信号蛋白的分布。在哺乳动物系统中，我们确定了趋化因子受体 CXCR1 在单细胞水平上的空间分布并可视化了信号级联。在 D. discoideum 系统中，我们监测了趋化细胞膜中 GPCR 的时间和空间活动。检测到 G 蛋白亚基之间的相互作用以及 GPCR 诱导的单细胞中 G 蛋白的解离。