Hydroxynonenal induces glutathione synthesis through JNK

羟基壬烯醛通过 JNK 诱导谷胱甘肽合成

基本信息

批准号：
7878839
负责人：
HENRY Jay FORMAN
金额：
$ 32.15万
依托单位：
UNIVERSITY OF CALIFORNIA, MERCED
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-09-22 至 2011-12-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7878839
关键词：
4 hydroxynonenal Address Affinity Chromatography Air Pollutants Anabolism Antibodies Antioxidants Be++ element Beryllium Binding Biological Assay Catalytic Domain Cell Extracts Cells Chromatin Complex DNA Dominant-Negative Mutation Elements Enzyme Induction Enzymes Epithelial Cells Excision Exposure to FarGo GCLC gene GCLM gene Gene Expression Genes Genetic Transcription Glutamate-Cysteine Ligase Glutathione Goals Hour Human Hydrogen Peroxide Immunoprecipitation In Situ Indium Inflammation Injury Interphase Cell Investigation Ligase Gene Lipid Peroxidation Luciferases Lung Lung diseases MAPK14 gene MAPK8 gene Mass Spectrum Analysis Measurement Measures Membrane Metabolism Nuclear Protein Nuclear Proteins Oxidants Oxidative Stress Pathologic Processes Pathway interactions Peptides Phosphotransferases Physiological Post-Translational Protein Processing Proteins Regulation Reporter Research Personnel Rest Reverse Transcriptase Polymerase Chain Reaction Role Signal Pathway Signal Transduction Signaling Protein Stress System Tissues Transcription Factor AP-1 Transcription factor genes Western Blotting Xenobiotics chromatin immunoprecipitation design human GCLC protein inhibitor/antagonist lung injury overexpression oxidation oxidative damage programs protein folding response stress activated protein kinase transcription factor

项目摘要

DESCRIPTION (provided by applicant): Our long range goal is to understand how to regulate glutathione (GSH) biosynthesis and thereby protect lungs from oxidative damage. Adaptation to oxidative stress in the lung involves an increase in the ability of cells to remove reactive and toxic molecules. Perhaps the most common mechanism for this is an increase in the ability to synthesize the endogenous antioxidant, GSH, which is essential for adaptation as it is critical in removal of hydroperoxides and toxic lipid peroxidation products, such as 4-hydroxynonenal (HNE). HNE is produced during exposure to any kind of oxidative stress but, interestingly, is one of the strongest inducers of GSH synthesis through increased transcription of both the regulatory and catalytic subunits of glutamate cysteine ligase (GCL). GCL catalyzes the rate limiting and first step in GSH synthesis. There is much evidence supporting roles for TRE and EpRE cis elements in regulation of the two GCL genes. We found that a little explored mode of activation of the EpRE and TRE elements is transcription factor (TF) switching. Our results also indicated that inhibition of the stress activated protein kinase, JNK, results in complete suppression of GCL induction by HNE. Thus, we hypothesize that HNE induced transcription of the two GCL genes involves switching from inactive or suppressing EpRE and TRE TF complexes to transcriptionally active complexes. We also hypothesize that HNE activation of the JNK signaling pathway is critical to both EpRE and AP-1 activation in GCL gene expression. The aims are: 1) to determine how the changes in the transcription factor binding complexes that bind to EpRE and TRE binding complexes in response to HNE cause increased transcription of both GCL genes and 2) to determine the mechanism of JNK activation by HNE. Both normal human bronchial epithelial cells (NHBE) and HBE1 cells will be used in exposures to subtoxic concentrations of HNE. In Aim 1, we will identify potential TFs by DNA affinity chromatography and LC-MS/MS analysis and then use chromatin immunoprecipitation (ChiP) assays to identify the EpRE and TRE TF complexes in the context of the whole GCL genes in situ. "NoShift" and Shift-Western assays will be used to quantify changes in TF binding and TF functionality will be determined by silencing TF genes and measuring EpRE and TRE driven luciferase reporters. In Aim 2, the effect of JNK inhibition on binding and function of TFs bound to EpRE will be examined along with a determination of which protein in the JNK pathway binds HNE. Relevance: Oxidative damage is a major component of lung injury during inflammation, other respiratory diseases and in exposure to air pollutants. The endogenous antioxidant glutathione increases during adaptation to sublethal oxidative stress through induction of the enzyme, GCL. In this investigation, the mechanism through which GCL increases in response to a toxic product of membrane oxidation will be investigated, hopefully leading to understanding of how to increase GSH without the use of toxic agents.

描述（由申请人提供）：我们的远距离目标是了解如何调节谷胱甘肽（GSH）生物合成，从而保护肺部免受氧化损伤。对肺中氧化应激的适应性涉及细胞去除反应性和有毒分子的能力。这样做的最常见机制也许是增强了内源性抗氧化剂GSH的能力，这对于适应至关重要，因为它对于去除氢过氧化物和有毒的脂质过氧化产物，例如4-羟基硝基（HNE）至关重要。 HNE是在暴露于任何类型的氧化应激期间产生的，但有趣的是，通过增加谷氨酸半胱氨酸连接酶（GCL）的调节和催化亚基的转录，是GSH合成的最强诱导剂之一。 GCL催化GSH合成的速率限制和第一步。有很多证据支持TRE和EPRE CIS元素在调节两个GCL基因中的作用。我们发现，EPRE和TRE元素激活的一种少量探索方式是转录因子（TF）切换。我们的结果还表明，抑制应激激活的蛋白激酶JNK，导致HNE完全抑制GCL诱导。因此，我们假设HNE诱导的两个GCL基因的转录涉及从非活性或抑制EPRE和TRE TF复合物转换为转录活性复合物。我们还假设JNK信号通路的HNE激活对于GCL基因表达中的EPRE和AP-1激活至关重要。目的是：1）确定转录因子结合复合物的变化如何与EPRE和TRE结合复合物响应响应HNE的响应导致GCL基因的转录增加和2）确定HNE通过HNE激活JNK激活的机理。正常的人支气管上皮细胞（NHBE）和HBE1细胞都将用于暴露于无毒性浓度的HNE。在AIM 1中，我们将通过DNA亲和力色谱和LC-MS/MS分析来鉴定潜在的TF，然后使用染色质免疫沉淀（CHIP）测定法在整个GCL基因的情况下鉴定EPRE和TRE TF复合物。 “ Noshift”和Shift-Western分析将用于量化TF结合的变化，而TF功能将通过沉默的TF基因并测量EPRE和TRE驱动的荧光素酶报道器来确定。在AIM 2中，将检查JNK抑制对与EPRE结合的结合和功能的影响，并确定JNK途径中哪种蛋白质结合HNE的蛋白质。相关性：氧化损伤是炎症期间肺损伤，其他呼吸道疾病和暴露于空气污染物的主要组成部分。内源性抗氧化剂谷胱甘肽通过诱导酶GCl的诱导在适应性氧化应激期间增加。在这项研究中，将研究GCL响应膜氧化有毒产物的机制，希望能够理解如何在不使用有毒剂的情况下增加GSH。

项目成果

期刊论文数量（65）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Glutathione synthesis and its role in redox signaling.

DOI：
10.1016/j.semcdb.2012.03.017
发表时间：
2012-09
期刊：
SEMINARS IN CELL & DEVELOPMENTAL BIOLOGY
影响因子：
7.3
作者：
Zhang, Hongqiao;Forman, Henry Jay
通讯作者：
Forman, Henry Jay

C-Myc is a Nrf2-interacting protein that negatively regulates phase II genes through their electrophile responsive elements.

DOI：
10.1002/iub.314
发表时间：
2010-03
期刊：
IUBMB LIFE
影响因子：
4.6
作者：
Levy, Smadar;Forman, Henry Jay
通讯作者：
Forman, Henry Jay

A549 subclones demonstrate heterogeneity in toxicological sensitivity and antioxidant profile.

A549 亚克隆在毒理学敏感性和抗氧化特性方面表现出异质性。

DOI：
10.1152/ajplung.00025.2002
发表时间：
2002
期刊：
American journal of physiology. Lung cellular and molecular physiology
影响因子：
0
作者：
Watanabe,Nobuo;Dickinson,DaleA;Krzywanski,DavidM;Iles,KarenE;Zhang,Hongqiao;Venglarik,CharlesJ;Forman,HenryJay
通讯作者：
Forman,HenryJay

Hyperthermic stress-induced increase in the expression of glutamate-cysteine ligase and glutathione levels in the symbiotic sea anemone Aiptasia pallida.

高温应激诱导共生海葵 Aiptasia pallida 中谷氨酸-半胱氨酸连接酶表达和谷胱甘肽水平增加。

DOI：
10.1016/j.cbpb.2008.06.007
发表时间：
2008
期刊：
Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology
影响因子：
0
作者：
Sunagawa,Shinichi;Choi,Jinah;Forman,HenryJay;Medina,Mónica
通讯作者：
Medina,Mónica

Extracellular glutathione and gamma-glutamyl transpeptidase prevent H2O2-induced injury by 2,3-dimethoxy-1,4-naphthoquinone.