Integration of Inflammatory Signaling and the Unfolded Protein Response by Nitrosylation Signaling in Obesity

肥胖中炎症信号传导与亚硝基化信号传导的未折叠蛋白反应的整合

基本信息

批准号：
10062953
负责人：
Ling Yang
金额：
$ 38.13万
依托单位：
UNIVERSITY OF IOWA
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-12-01 至 2022-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10062953
关键词：
Address Affect Animal Model Attenuated Binding Proteins Biochemical Biological Butyric Acids Cells Chemicals Chronic Data Development Diabetes Mellitus Dyslipidemias Endoplasmic Reticulum Enhancers Enzymes Excision Failure Functional disorder Glutathione Goals Hepatic Homeostasis Human Immunoprecipitation Impairment Inflammation Inflammatory Inositol Insulin Insulin Resistance Knockout Mice Knowledge Laboratory Research Liver Maps Mediating Mediator of activation protein Metabolic Metabolic Diseases Metabolism Mission Modification Molecular Mus NOS2A gene Nicotinamide adenine dinucleotide Nitric Oxide Nitric Oxide Synthase Non-Insulin-Dependent Diabetes Mellitus Obese Mice Obesity Organelles Outcome Output Oxidoreductase Pancreatic ribonuclease Pathology Pathway interactions Post-Translational Protein Processing Production Protein S Proteins Proteome Proteomics Public Health RNA Processing RNA Splicing RNA-Protein Interaction Regulation Research Resistance Ribonucleases Role S-Nitrosoglutathione Signal Transduction Site Speed Stress Supplementation Testing Therapeutic Thinness Tissues United States National Institutes of Health Up-Regulation base biological adaptation to stress blood glucose regulation crosslink diabetic patient diet-induced obesity disability endoplasmic reticulum stress improved in vivo innovation insight insulin sensitivity mouse S-nitrosoglutathione reductase mouse model new therapeutic target nitrosative stress novel obesity development overexpression protein complex response

项目摘要

PROJECT SUMMARY In the setting of obesity, endoplasmic reticulum (ER) stress has been identified as a prominent feature in metabolic tissues in both animal models and in humans. To cope with ER stress, cells activate the unfolded protein response (UPR) to mitigate stress. However, failure of the UPR results in chronic unresolved stress, contributing to the development of obesity-induced insulin resistance. Nitric oxide (NO) is a key mediator of obesity-associated inflammation. We recently demonstrated that NO-mediated protein modification (S- nitrosylation) impairs the RNase activity of inositol-requiring enzyme-α (IRE1α), resulting in unresolved ER stress and insulin resistance. Although nitric oxide synthase (NOS) provides the general intracellular NO pools for protein S-nitrosylation, the rate of these modifications is also affected by the targeted removal of NO groups by protein denitrosylation. Our strong preliminary data demonstrate that obesity impairs the activity of S- nitrosoglutathione reductase (GSNOR, a major denitrosylase), leading to elevated nitrosative stress in the liver. Furthermore, deletion or overexpression of GSNOR directly regulates the S-nitrosylation state of IRE1α and ER function in mice with diet-induced obesity (DIO). Notably, liver-specific GSNOR overexpression ameliorated obesity-associated insulin resistance. However, the molecular mechanism that underlies the regulation of ER homeostasis by GSNOR-mediated denitrosylation signaling is unknown. We propose to test the central hypothesis that obesity attenuates GSNOR-dependent protein denitrosylation, resulting in elevated nitrosative stress in the ER that contributes to obesity-associated hepatic insulin resistance. To test this hypothesis, we will undertake 2 specific aims. In Aim 1, we will: 1) determine whether liver-specific GSNOR deletion impacts hepatic insulin sensitivity and whole body glucose homeostasis using DIO mouse model; 2) establish whether GSNOR regulates hepatic insulin action directly via modulation of UPR; and 3) assess the therapeutic potential of enhancing hepatic GSNOR activity in obese mice by glutathione supplementation and enhancing cellular nicotinamide adenine dinucleotide (NAD) metabolism. In Aim 2, we will: 1) profile the ER S-nitrosylation proteome and characterize the endogenous S-nitrosylation sites on IRE1α; 2) address how GSNOR-mediated denitrosylation signaling modulates the IRE1α RNase activity; and 3) establish how GSNOR-mediated denitrosylation signaling affects IRE1α interactome formation. The approaches used here are innovative, combining the use of cellular and molecular biological analysis, biochemical analysis for S-nitrosylated proteins, elucidating the ER S-nitrosylation proteome, protein-RNA interaction analysis, and in vivo mouse metabolic profiling. The mechanisms elucidated in this project will provide further understanding of how inflammatory and ER stress pathways are integrated in the context of obesity-associated hepatic insulin resistance, dyslipidemia and ultimately type 2 diabetes. Insight into the mechanisms by which maladaptive organelle stress responses drive these pathologies should speed development of novel therapeutic targets for metabolic diseases.

项目概要在肥胖的情况下，内质网（ER）应激已被确定为肥胖的一个突出特征。动物模型和人类的代谢组织为了应对内质网应激，细胞激活未折叠的细胞。蛋白质反应（UPR）来缓解压力然而，UPR的失败会导致长期未解决的压力，一氧化氮（NO）是导致肥胖引起的胰岛素抵抗的关键介质。我们最近证明了 NO 介导的蛋白质修饰（S-）。亚硝基化）损害肌醇需要酶-α (IRE1α) 的 RNase 活性，导致 ER 无法解决尽管一氧化氮合酶 (NOS) 提供了一般的细胞内 NO 池。对于蛋白质 S-亚硝基化，这些修饰的速率也受到 NO 基团靶向去除的影响我们强有力的初步数据表明，肥胖会损害 S- 的活性。亚硝基谷胱甘肽还原酶（GSNOR，一种主要的脱亚硝基酶），导致肝脏亚硝化应激升高。此外，GSNOR 的缺失或过表达直接调节 IRE1α 的 S-亚硝基化状态，饮食诱导肥胖 (DIO) 小鼠的 ER 功能显着改善，肝脏特异性 GSNOR 过度表达得到改善。然而，肥胖相关的胰岛素抵抗的分子机制。 GSNOR 介导的去亚硝基化信号传导的稳态尚不清楚，我们建议测试中枢。假设肥胖会减弱 GSNOR 依赖性蛋白质去亚硝基化，导致亚硝化水平升高内质网应激会导致肥胖相关的肝脏胰岛素抵抗。为了检验这一假设，我们进行了研究。将实现 2 个具体目标在目标 1 中，我们将： 1) 确定肝脏特异性 GSNOR 缺失是否会产生影响。使用DIO小鼠模型2)建立肝脏胰岛素敏感性和全身葡萄糖稳态； GSNOR 通过调节 UPR 直接调节肝脏胰岛素作用；3) 评估治疗潜力；通过补充谷胱甘肽增强肥胖小鼠肝脏 GSNOR 活性和增强细胞在目标 2 中，我们将：1) 分析 ER S-亚硝基化。蛋白质组并表征 IRE1α 上的内源 S-亚硝基化位点 2) 解决 GSNOR 介导的问题；去亚硝基化信号调节 IRE1α RNase 活性；3) 确定 GSNOR 是如何介导的；去亚硝基化信号传导影响 IRE1α 相互作用组的形成。这里使用的方法是创新的。结合细胞和分子生物学分析、S-亚硝基化蛋白质的生化分析，阐明 ER S-亚硝基化蛋白质组、蛋白质-RNA 相互作用分析和小鼠体内代谢该项目阐明的机制将有助于进一步了解炎症和炎症如何发生。内质网应激途径与肥胖相关的肝脏胰岛素抵抗、血脂异常相关以及最终的 2 型糖尿病的深入了解适应不良细胞器应激反应的机制。驱动这些病理学应该加速代谢疾病新治疗靶点的开发。