Regulation of Chemotactic Signaling

趋化信号的调节

• 批准号: 10386438
• 负责人: Miho Iijima
• 金额: $ 12.62万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10386438
• 关键词: Animal Model; Arthritis; Asthma; Atherosclerosis; Behavior; Biochemical; Cell Line; Cell Nucleus; Cell membrane; Cells; Cellular Structures; Chemicals; Chemotaxis; Detection; Disease; Embryonic Development; Fluorescence; Funding; Funding Opportunities; GTP Binding; Goals; High Pressure Liquid Chromatography; Human; Immune response; In Vitro; Knock-out; Malignant Neoplasms; PTEN gene; Phosphorylation; Physiological Processes; Proto-Oncogene Proteins c-akt; Race; Regulation; Signal Transduction; System; Therapeutic Intervention; X-Ray Crystallography; cell motility; extracellular; in vivo; live cell imaging; migration; parent grant; protein complex; protein protein interaction; reconstitution; single molecule; therapeutic development; tumorigenesis; wound healing

Summary Through the funding opportunity described in PAR-20-272, this proposal seeks funds to purchase a fluorescence-detection HPLC (high-performance liquid chromatography) system to quantitatively assess stimulation-induced dynamic assembly of protein complexes in chemotactic signaling. The goals of the parent grant R35GM131768 are to investigate how cells sense extracellular chemical gradients and control their migration behaviors. First, we will determine how phosphorylated, GDP-bound RacE activates TORC2 to facilitate AKT phosphorylation at the leading end of migrating cells while GTP-bound RacE inhibits TORC2 at the trailing end. This will involve multiple experimental approaches, including cellular reconstitution using knockout cell lines, biochemical reconstitution of RacE-regulated TORC2 activity with purified proteins, protein interaction analyses at the single-molecule level in vitro, live imaging of cells, and structural analysis using X-ray crystallography. Second, we will decipher a key mechanism controlling human PTEN localization at the plasma membrane and nucleus. Third, we will determine the effects of the manipulation of PTEN localization on tumorigenesis in vivo using animal models.

概括 通过 PAR-20-272 中描述的资助机会，本提案寻求资金 购买荧光检测 HPLC（高效液相色谱）系统 定量评估刺激诱导的蛋白质复合物动态组装 趋化信号传导。母基金 R35GM131768 的目标是研究细胞如何 感知细胞外化学梯度并控制其迁移行为。首先，我们将 确定磷酸化、GDP 结合的 RacE 如何激活 TORC2 以促进 AKT 迁移细胞前端的磷酸化，而 GTP 结合的 RacE 抑制 TORC2 尾端。这将涉及多种实验方法，包括细胞 使用敲除细胞系重建，RacE 调节的 TORC2 生化重建 纯化蛋白质的活性，体外单分子水平的蛋白质相互作用分析， 细胞实时成像，以及使用 X 射线晶体学进行结构分析。其次，我们将 破译控制人类 PTEN 在质膜上定位的关键机制 核。第三，我们将确定 PTEN 定位的操纵对 使用动物模型进行体内肿瘤发生。