Defining Molecular Determinants of Type 2 Diabetes Using Quantitative Proteomics

使用定量蛋白质组学定义 2 型糖尿病的分子决定因素

• 批准号: 8335569
• 负责人: Paul A. Grimsrud
• 金额: $ 5.57万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8335569
• 关键词: 4 hydroxynonenal; Affect; Aldehydes; All-Trans-Retinol; Analytical Chemistry; Antioxidants; Attenuated; BTBR Mouse; Basic Science; Binding; Biochemical; Biochemistry; Biogenesis; Biological; Biological Assay; Biology; Cell Culture Techniques; Cell physiology; Cells; Cellular biology; Collection; Control Animal; Data; Data Set; Development; Diabetes Mellitus; Disease; Doctor of Philosophy; Elongation Factor; Environment; Enzymes; Epidemic; Epitopes; Family member; Functional disorder; Genetic; Goals; Immunoprecipitation; In Vitro; Incidence; Inflammatory; Insulin Resistance; Investigation; Laboratories; Link; Lipid Peroxidation; Liver; Liver Mitochondria; Mass Spectrum Analysis; Metabolic; Metabolic Diseases; Metabolism; Minnesota; Mitochondria; Mitochondrial Proteins; Modeling; Molecular; Monitor; Mus; Mutate; Non-Insulin-Dependent Diabetes Mellitus; Obesity; Oxidation-Reduction; Oxidoreductase; Peripheral; Phosphorylation; Phosphorylation Site; Play; Postdoctoral Fellow; Predisposition; Production; Protein Analysis; Protein Isoforms; Proteins; Proteome; Proteomics; RNA; RNA Interference; Reaction; Recombinant Proteins; Recombinants; Research; Research Project Grants; Resistance; Retinal; Retinoids; Role; Selenocysteine; Signal Transduction; Site-Directed Mutagenesis; Surveys; Testing; Therapeutic Intervention; Tissues; Training; Translations; United States; Universities; Wisconsin; Work; aldehyde dehydrogenases; base; cohort; diabetic; genetic regulatory protein; in vitro activity; insight; instrumentation; member; mitochondrial dysfunction; mutant; novel; overexpression; oxidized lipid; research study; resistant strain; response; selenoprotein; stoichiometry

The objective of the proposed research is to determine how alterations in mitochondrial protein abundance and phosphorylation contribute to Type 2 Diabetes Mellitus (T2DM). Recent studies have linked mitochondrial dysfunction to insulin resistance. Dr. Alan Attie, co-sponsor of this proposal, uses genetics to understand the propensity to develop T2DM. Dr. David Pagliarini, sponsor of this proposal, focuses on the contribution of mitochondrial dysfunction to various diseases and understanding the basic science of mitochondrial biogenesis. Dr. Joshua Coon, the sponsor of my original application and a close collaborator on this proposal, is a leader in the development and application of mass spectrometry (MS) instrumentation for protein analysis. Having transition from the Coon group to start a second post-doc in the Pagliarini lab, I am continuing this project as the leading member of our interdisciplinary collaborative team. I have completed collection of an exhaustive quantitative proteomics dataset that constituted the bulk of my proposed research for this project in my original application. I observed highly reproducible alterations in protein abundance and phosphorylation in liver mitochondria in a cohort of over forty mice, which are either susceptible (B6) or resistant (BTBR) to developing T2DM when made obese. I am currently using this proteomics screen as preliminary data for hypotheses-driven targeted biological investigation. The rationale for the proposed research is that determining how phosphorylation modulates redox-regulatory proteins in obesity will open new avenues for developing therapeutic interventions for type 2 diabetes. The co-sponsoring environment will help facilitate my goal of starting an independent academic laboratory focused on utilizing proteomics and targeted biology to study redox signaling in metabolic disease. I will employ both biochemical, cell biology, and targeted proteomic approaches to carrying out the following Aims: Aim 1. Determine how the obesity-induced phosphorylation of the selenocysteine-specific elongation factor (Eefsec) regulates the production of redox-regulatory selenoproteins. I will test the hypothesis that enhanced obesity-induced phosphorylation of the non-mitochondrial isoform of Eefsec in diabetes- resistant B6 mice attenuates the production of inflammatory secreted selenoproteins without decreasing levels of key mitochondrial antioxidants, which are regulated by a mitochondrial-localized Eefsec isoform. Aim 2. Elucidate the role of obesity-induced phosphorylation of mitochondrial redox enzymes in regulating the abundance of bioactive lipophilic aldehydes. I will test the hypothesis that obesity-induced phosphorylation of dehydrogenase/reductase SDR family member 4 (Dhrs4) alters the levels of retinoid metabolites in diabetic BTBR mice through regulating the enzyme's retinal reducing activity. A parallel hypothesis I will test is that induction of aldehyde dehydrogenase 3A2 (Aldh3a2) expression and phosphorylation is a compensatory response in obesity for detoxifying reactive lipid-peroxidation products.

拟议研究的目的是确定线粒体蛋白丰度和磷酸化的改变如何有助于2型糖尿病（T2DM）。最近的研究将线粒体功能障碍与胰岛素抵抗联系起来。该提案的共同发起人艾伦·阿蒂（Alan Attie）博士使用遗传学来了解发展T2DM的倾向。该提案的赞助商David Pagliarini博士的重点是线粒体功能障碍对各种疾病的贡献，并了解线粒体生物发生的基础科学。我原始应用程序的赞助商Joshua Coon博士是该提案的密切合作者，是质谱（MS）仪器进行蛋白质分析的开发和应用的领导者。从Coon Group过渡到在Pagliarini实验室开始了第二个大赛后，我将继续作为我们跨学科合作团队的主要成员。 我已经完成了一个详尽的定量蛋白质组学数据集的集合，该数据集构成了我在原始应用中对该项目的大部分研究。我观察到蛋白质丰度和磷酸化的高度可再现的改变，在一组超过四只小鼠的队列中，它们易感（B6）或耐药性（BTBR）在肥胖时会发育T2DM。我目前正在使用此蛋白质组学屏幕作为假设驱动的靶向生物学研究的初步数据。拟议研究的基本原理是确定肥胖症中磷酸化如何调节氧化还原调节蛋白将开辟新的途径，以开发针对2型糖尿病的治疗干预措施。共同赞助的环境将有助于促进我的目标，即启动专注于利用蛋白质组学和靶向生物学的独立学术实验室来研究代谢疾病中的氧化还原信号。我将采用生化，细胞生物学和靶向蛋白质组学方法来执行以下目的： 目的1。确定肥胖诱导的硒雌激素特异性伸长因子（EEFSEC）的磷酸化如何调节氧化还原调节性硒蛋白的产生。我将测试以下假设：增强肥胖性诱导的EEFSEC非细胞软骨同工型在糖尿病 - 耐药的B6小鼠中可减弱炎症性硒蛋白的产生，而不会降低关键水平的关键抗氧化剂的水平，而这些抗氧化剂的水平降低了，这些抗氧化剂是通过粒子质量质量构造的。 目标2。阐明肥胖诱导的线粒体氧化还原酶磷酸化在调节丰富的生物活性亲脂醛中的作用。我将测试以下假设：肥胖诱导的脱氢酶/还原酶SDR家族成员4（DHRS4）通过调节酶的视网膜减少活性来改变糖尿病BTBR小鼠中类视丁代谢物的水平。一个平行的假设I将检验，是诱导醛脱氢酶3A2（ALDH3A2）的表达，磷酸化是肥胖的补偿性反应，用于解毒反应性脂质过氧产物。