Redox Regulation of Intracellular Calcium Signaling

细胞内钙信号传导的氧化还原调节

基本信息

批准号：
8905057
负责人：
Gyorgy Hajnoczky
金额：
$ 35.04万
依托单位：
THOMAS JEFFERSON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-04-01 至 2019-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8905057
关键词：
Address Affect Agonist Antibodies Apoptosis Arrhythmia Bile fluid Binding Biochemical Biological Assay C-terminal Calcium Calcium Signaling Cardiovascular Diseases Cell Death Cell Fractionation Cell physiology Cells Cellular Stress Collaborations Cysteine Data Defect Dependence Diabetes Mellitus Disease Drug Targeting Drug or chemical Tissue Distribution Endoplasmic Reticulum Experimental Models Fluorescence Frequencies Fructose Gene Expression Genetic Transcription Glutathione Disulfide Goals Growth Factor Heart Diseases Heart Hypertrophy Homo Hormones Hydrogen Peroxide ITPR1 gene Image Incubated Inositol Ion Channel Life Ligand Binding Domain Ligands Liver Location Malignant Neoplasms Maps Mass Spectrum Analysis Measurement Measures Mediating Membrane Metabolism Methods Microinjections Mitochondria Modeling Modification Molecular Mutate N-terminal NADP NADPH Oxidase Neurodegenerative Disorders Neurotransmitters Non-Insulin-Dependent Diabetes Mellitus Oxidants Oxidation-Reduction Oxidative Stress Pathway interactions Pharmaceutical Preparations Play Production Property Protein Isoforms Proteins Proteome Reaction Regulation Role Signal Transduction Sirolimus Source Spinocerebellar Ataxias Stress Sulfhydryl Compounds System Tacrolimus Binding Proteins Testing Thimerosal Tissues Transmembrane Domain Vaccines Work base catalase cell motility cell type design endoplasmic reticulum stress extracellular imaging modality in vivo mutant novel novel therapeutics palmitoylation public health relevance receptor reconstitution research study response

项目摘要

DESCRIPTION (provided by applicant): An elevation of cytosolic free calcium concentration is an integral component of the mechanism by which cells respond to hormones, growth factors and neurotransmitters. D-myo-inositol 1,4,5-trisphosphate ( IP3 ) is an intracellular messenger mediating the mobilization of Ca2+ from intracellular stores by interaction with an ubiquitous receptor ( IP3R ) that acts as a ligand-gated Ca2+ channel. IP3Rs are redox sensitive channels and are sensitized by oxidative stress. However, the molecular basis of this regulation is poorly understood. Ca2+ released from IP3Rs is locally transmitted to the mitochondria and can stimulate metabolism, and in higher amounts, can also initiate cell death. The overarching hypothesis of this study is that redox modulation of IP3Rs is an important component of the regulation of Ca2+ signals in cell death pathways. The proposal encompasses the following three specific aims: 1] To measure and map redox changes in IP3Rs. We have developed methods to determine the redox state of IP3Rs in vivo which will be used to quantitate the effects of exogenous and endogenous agents causing oxidative stress. Preliminary studies using mass-spectroscopy identify a subset of 11 cysteines that become oxidized in IP3R-1. The type of oxidative modifications occurring will be identified. Redox-sensitive thiols will be mutate and the functional sensitivity to oxidative stress will be assessed. 2] To measure IP3R redox state at the ER/mitochondrial junction. We will test the hypothesis that the pool of IP3Rs located at the ER/mito junction is particularly prone to ROS modifications. We will employ subcellular fractionation and imaging methods utilizing targeted IP3Rs, ROS-sensitive fluorescent proteins, ROS-producing photosensitive probes and targeted catalases. 3] To investigate the role of IP3R redox changes in models of ER stress/apoptosis. We will test the hypothesis that ER-resident NADPH oxidases play an important role in IP3R redox regulation. Liver will be used as an experimental model to induce ER stress. The role of IP3R redox regulation in ER stress pathways activated by fructose will be examined. The long-term goal of the proposal is to obtain a detailed understanding of how oxidative stress impacts intracellular Ca2+ signaling under normal and disease conditions.

描述（由申请人提供）：胞质游离钙浓度的升高是细胞对激素、生长因子和神经递质作出反应的机制的一个组成部分。D-肌醇 1,4,5-三磷酸 (IP3) 是一种机制。细胞内信使通过与作为配体门控 Ca2+ 通道的普遍存在的受体 (IP3R) 相互作用，介导细胞内储存的 Ca2+ 的动员。 IP3R 是氧化还原敏感通道，对氧化应激敏感，但人们对这种调节的分子基础知之甚少。本研究的总体假设是 IP3R 的氧化还原调节是细胞死亡途径中 Ca2+ 信号调节的重要组成部分。该提案涵盖以下三个具体目标： 1] 测量和确定。我们开发了确定 IP3R 体内氧化还原状态的方法，该方法将用于定量引起氧化应激的外源性和内源性物质的影响，并使用质谱法鉴定了 11 个半胱氨酸的子集。 IP3R-1 中发生的氧化修饰的类型将发生突变，并且对氧化还原敏感的硫醇的功能敏感性将被识别。 2] 为了测量 ER/线粒体连接处的 IP3R 氧化还原状态，我们将测试位于 ER/线粒体连接处的 IP3R 库特别容易发生 ROS 修饰的假设。利用靶向 IP3R、ROS 敏感荧光蛋白、产生 ROS 的光敏探针和靶向过氧化氢酶的成像方法 3] 研究 IP3R 氧化还原变化在模型中的作用。内质网应激/细胞凋亡。我们将测试内质网驻留的 NADPH 氧化酶在 IP3R 氧化还原调节中发挥重要作用的假设，将使用肝脏作为诱导内质网应激的实验模型来验证 IP3R 氧化还原调节在内质网应激途径中的作用。该提案的长期目标是详细了解氧化应激如何影响正常和疾病条件下的细胞内 Ca2+ 信号传导。