Regulation of intracellular calcium signaling and phosphoinositide metabolism by IRBIT

IRBIT 对细胞内钙信号传导和磷酸肌醇代谢的调节

• 批准号: 10702168
• 负责人: Martin Kruse
• 金额: $ 41.54万
• 依托单位: BATES COLLEGE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-07 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10702168
• 关键词: Action Potentials; Affect; Alzheimer' s Disease; Binding Proteins; Calcium; Calcium Signaling; Calcium ion; Candidate Disease Gene; Catalytic Domain; Cell Line; Cells; Confocal Microscopy; Critical Pathways; Cytoplasm; Data; Disease; Embryo; Endoplasmic Reticulum; Epilepsy; Family; Gene Expression; Generations; Genes; Health; Human; ITPR1 gene; Image; Inositol; Ions; Kidney; Kinetics; Knowledge; Lipid Binding; Lipids; Mammalian Cell; Mass Spectrum Analysis; Mediating; Metabolism; Minor; Molecular; Nature; Neurotransmitters; Organelles; Parkinson Disease; Pathologic; Patients; Persons; Phosphatidylinositol Phosphates; Phosphatidylinositols; Phospholipids; Phosphotransferases; Process; Proteins; Publications; Purinoceptor; RNA; Regulation; Research; Research Personnel; Research Technics; Role; Severities; Signal Pathway; Signal Transduction; Small Interfering RNA; Students; System; United States; Western Blotting; antagonist; confocal imaging; experience; experimental study; gene product; improved; inorganic phosphate; insight; kidney cell; knock-down; meetings; nervous system disorder; neuropathology; novel; novel therapeutic intervention; overexpression; transcriptome sequencing; treatment strategy; whole genome

Project Summary Epilepsy and Alzheimer’s disease affect more than nine million people in the United States. Despite their severity and decades of research, we still lack an understanding of the molecular mechanisms governing these diseases and other related neuropathological conditions. We propose that a recently discovered protein, IRBIT (IP3R binding protein released with inositol 1,4,5-trisphosphate), functions as a regulator of two major signaling pathways, lipid-based phosphoinositide signaling and release of calcium ions from intracellular organelles into the cytoplasm, that have both been shown to be critically involved in these neurological diseases and other pathological conditions. Based on this hypothesis, we will characterize how IRBIT regulates these signaling pathways by (i) characterizing calcium signaling in wild-type and IRBIT-/- human embryonic kidney (HEK293) cells, (ii) utilizing these cell lines to analyze phosphoinositide composition by lipid mass spectrometry and determine the kinetics of phosphoinositide metabolism using fluorescent phosphoinositide-binding proteins in live- cell confocal imaging experiments, and (iii) using an unbiased whole-genome expression analysis via RNA-Seq in wild-type and IRBIT-/- human embryonic kidney cells to assess how IRBIT’s activity influences expression of genes involved in these two signaling networks. The data from the proposed experiments will allow us to gain insight into a novel mechanism by which one protein, IRBIT, regulates two signaling pathways that are of critical importance in practically all mammalian cells. This enhanced understanding of these signaling pathways and their regulator mechanisms has the potential to further our knowledge about the molecular nature of debilitating diseases such as epilepsy and Alzheimer’s and allow us to develop improved treatment strategies for patients affected by these conditions.

项目概要 癫痫和阿尔茨海默病影响着美国超过 900 万人。尽管 尽管其严重性和数十年的研究，我们仍然缺乏对其分子机制的了解 控制这些疾病和其他相关的神经病理学病症。我们建议最近 发现蛋白质 IRBIT（与肌醇 1,4,5-三磷酸一起释放的 IP3R 结合蛋白）的功能 作为两个主要信号传导途径的调节剂，基于脂质的磷酸肌醇信号传导和释放 钙离子从细胞内细胞器进入细胞质，这两者都被证明是 严重参与这些神经系统疾病和其他病理状况。基于此 假设，我们将通过 (i) 表征 IRBIT 如何调节这些信号通路 野生型和 IRBIT-/- 人胚肾 (HEK293) 细胞中的钙信号传导，(ii) 利用这些 细胞系通过脂质质谱分析磷酸肌醇成分并确定 使用荧光磷酸肌醇结合蛋白在活体中进行磷酸肌醇代谢动力学 细胞共聚焦成像实验，以及（iii）使用无偏全基因组表达分析 在野生型和 IRBIT-/- 人胚胎肾细胞中进行 RNA 测序，以评估 IRBIT 的活性 影响这两个信号网络中涉及的基因的表达。拟议的数据 实验将使我们能够深入了解一种新的机制，通过这种机制，一种蛋白质 IRBIT 调节对几乎所有哺乳动物细胞都至关重要的两条信号传导途径。 增强对这些信号通路及其调节机制的理解具有以下意义： 进一步了解衰弱性疾病的分子性质的潜力，例如 癫痫和阿尔茨海默氏症，使我们能够为受影响的患者制定改进的治疗策略 通过这些条件。