GLUCOCORTICOID RECEPTOR POST-TRANSLATIONAL MODIFICATIONS IN INSULIN RESISTANCE

胰岛素抵抗中的糖皮质激素受体翻译后修饰

• 批准号: 9980364
• 负责人: Clay F. Semenkovich
• 金额: $ 34.31万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9980364
• 关键词: AF2; Adipose tissue; Adrenal Glands; Alanine; Animal Model; Animals; Anti-Inflammatory Agents; Antibodies; Antisense Oligonucleotides; Area; Back; Binding; Blood Circulation; Bypass; Caring; Cell Line; Cells; Central obesity; Chromatin; Chronic Obstructive Airway Disease; Clinical Treatment; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Cushing Syndrome; DNA; DNA Binding Domain; Data; Development; Dexamethasone; Diabetes Mellitus; Diet; Disease; Enzymes; Epidemic; Etiology; Event; Exposure to; Fatty Liver; Fatty acid glycerol esters; Future; Gene Activation; Gene Expression; Genes; Genetic Transcription; Glucocorticoid Receptor; Glucocorticoids; Hepatic; Hepatocyte; Hormones; Human; Hydrocortisone; Hydroxysteroid Dehydrogenases; Hyperlipidemia; Hypertension; In Vitro; Inflammatory; Inflammatory Bowel Diseases; Insulin; Insulin Resistance; Investigation; Knock-in; Knock-in Mouse; Lead; Leptin; Ligand Binding Domain; Ligands; Light; Link; Lipids; Lipolysis; Liver; Mediating; Metabolic; Metabolic Diseases; Metabolic syndrome; Metabolism; Mus; Mutate; Mutation; Nuclear Hormone Receptors; Obese Mice; Obesity; Operative Surgical Procedures; Oxidoreductase; Patients; Pattern; Pharmaceutical Preparations; Phosphorylation; Physiological; Post-Translational Protein Processing; Protein Isoforms; Proteins; Receptor Signaling; Reporter; Repression; Rheumatoid Arthritis; Risk Factors; Rodent Model; Role; Sampling; Serine; Serum; Signal Transduction; Stable Isotope Labeling; Stains; Stress; Testing; Tissues; Transactivation; Transgenic Organisms; Triglyceride Metabolism; Triglycerides; Visceral; Weight Gain; base; chromatin immunoprecipitation; diabetogenic; effective therapy; glucocorticoid receptor alpha; hepatoma cell; improved; in vivo; inhibitor/antagonist; insulin sensitivity; insulin sensitizing drugs; knock-down; lipid metabolism; liver biopsy; mutant; novel; receptor; response; small hairpin RNA

Abstract: There is a global epidemic of metabolic syndrome, characterized by insulin resistance, hypertension, and central obesity. Although the etiology of metabolic syndrome is still an active area of investigation, one leading hypothesis is that it is caused by elevated glucocorticoid (GCs) levels in key insulin responsive tissues such as liver and fat. GCs are stress hormones produced by the adrenal gland. GCs bind to the GC receptor (GR) a nuclear hormone receptor that alters transcription of target genes by multiple mechanisms including direct binding to DNA (transactivation) as well as protein-protein mediated repression. GCs are also effective therapies for many inflammatory diseases including chronic obstructive pulmonary disease, inflammatory bowel disease and rheumatoid arthritis, but their use is limited due the adverse metabolic effects including weight gain, insulin resistance and hypertension. Recently, several groups have identified GR post- translational modification (PTM) in cell lines, and these PTM events alter GR transcriptional activity. However, the role of GR PTM in physiologically relevant paradigms in vivo is unknown. Based on preliminary data, I hypothesize that GR PTM, specifically phosphorylation, mediates the complex effects of GC on the liver in vivo. This hypothesis will be tested in three Specific Aims: 1) We will determine the role of GR phosphorylation on hepatocyte metabolism and gene expression. This will be done by adding back wildtype or phosphomutant GR isoforms to GR knockdown hepatoma cells 2) We will determine the role of GR phosphorylation in mediating heaptic metabolism, gene expression and GR occupancy in the livers of mice. This will be done by analyzing metabolism, gene expression and chromatin immunoprecipitation with antibodies to GR. We have created novel GR knockin phosphomutant mice using CRISPRS. These S211A mice have a key phosphorylated serine residue mutated to alanine. We will determine which GC mediated changes in hepatic lipid metabolism, gene expression and GR occupancy are phosphorylation dependent. 3) We will determine if GR phosphorylation is associated with hepatic insulin resistance in samples from obese patients undergoing Roux-en-Y gastic bypass surgery. This will be done by staining liver biopsies from these patients (as well as controls) with phosphoGR specific antibodies. Currently selective GR modulators are being developed for the treatment of metabolic diseases as well as safer anti-inflammatory medications. Therefore, understanding the role of GR PTM in mediating effects of GCs should shed light on novel treatments for metabolic disease as well as common inflammatory conditions.

抽象的： 代谢综合征在全球范围内流行，其特征是胰岛素抵抗、高血压和 向心性肥胖。尽管代谢综合征的病因学仍然是一个活跃的研究领域，但一个领先的研究领域 假设是它是由关键胰岛素反应组织中糖皮质激素 (GC) 水平升高引起的，例如 肝脏和脂肪。 GC 是肾上腺产生的应激激素。 GC 与 GC 受体 (GR) 结合 核激素受体，通过多种机制改变靶基因的转录，包括直接 与 DNA 结合（反式激活）以及蛋白质-蛋白质介导的抑制。 GC也很有效 治疗许多炎症性疾病，包括慢性阻塞性肺病、炎症 肠道疾病和类风湿性关节炎，但由于不利的代谢影响，它们的使用受到限制，包括 体重增加、胰岛素抵抗和高血压。最近，一些研究小组已经确定了 GR 后 细胞系中的翻译修饰 (PTM)，这些 PTM 事件会改变 GR 转录活性。然而， GR PTM 在体内生理相关范例中的作用尚不清楚。根据初步数据，我 假设 GR PTM，特别是磷酸化，介导 GC 对肝脏的复杂影响 体内。该假设将通过三个具体目标进行检验：1）我们将确定 GR 磷酸化的作用 肝细胞代谢和基因表达的影响。这将通过添加回野生型或磷酸突变体来完成 GR 亚型与 GR 敲除肝癌细胞 2) 我们将确定 GR 磷酸化在 介导小鼠肝脏中的肝代谢、基因表达和 GR 占据。这将由 使用 GR 抗体分析代谢、基因表达和染色质免疫沉淀。我们有 使用 CRISPRS 创建了新型 GR 敲入磷酸突变小鼠。这些 S211A 鼠标有一把钥匙 磷酸化丝氨酸残基突变为丙氨酸。我们将确定哪些 GC 介导了肝脏的变化 脂质代谢、基因表达和 GR 占据均依赖于磷酸化。 3）我们将确定是否 肥胖患者样本中 GR 磷酸化与肝脏胰岛素抵抗相关 Roux-en-Y胃绕道手术。这将通过对这些患者的肝活检（以及 对照）与磷酸GR特异性抗体。目前正在开发选择性 GR 调制器 治疗代谢性疾病以及更安全的抗炎药物。因此，了解 GR PTM 在 GC 介导作用中的作用应为代谢疾病的新疗法提供启示 以及常见的炎症状况。