Nutritional regulation of leptin production

瘦素产生的营养调节

• 批准号: 7782186
• 负责人: Susan K Fried
• 金额: $ 21.28万
• 依托单位: BOSTON MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-06 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7782186
• 关键词: 3' Untranslated Regions; 3T3-L1 Cells; 5' Untranslated Regions; A kinase anchoring protein; Abbreviations; Acute; Adenosine Monophosphate; Adipocytes; Adipose tissue; Adrenergic Agents; Adrenergic beta-Agonists; Affect; Anabolism; Binding; Binding Sites; Biological Assay; Biology; Body fat; Catecholamines; Cell model; Cells; Chronic; Complex; Cues; Cultured Cells; Data; Deletion Mutation; Detection; Dexamethasone; Elements; Endocrine; Endoplasmic Reticulum; Family; Genetic Translation; Goals; Growth Factor; Hormones; Human; Hyperinsulinism; Immunity; Immunoprecipitation; In Vitro; Insulin; Insulin Resistance; Interleukin-1; Isoproterenol; Knowledge; Label; Leptin; Link; Luciferases; MAP Kinase Gene; Macronutrients Nutrition; Mediating; Messenger RNA; Metabolic; Metabolism; Mitogen-Activated Protein Kinases; Modeling; Mus; Mutation; Nutrient; Nutritional; Nutritional status; Obesity; Osteogenesis; Pathway interactions; Phosphatidylinositols; Phosphorylation; Phosphotransferases; Physiological; Polyribosomes; Production; Protein Binding; Protein Kinase; Protein Overexpression; Proteins; RNA-Binding Proteins; Rattus; Regulation; Relative (related person); Reporter; Reporter Genes; Reproduction; Research Personnel; Reverse Transcriptase Polymerase Chain Reaction; Ribonucleoproteins; Role; Signal Transduction; Starvation; Stress; System; TNF gene; Testing; Trans-Activators; Transfection; Translations; Tumor Necrosis Factor-alpha; Tumor Necrosis Factors; Untranslated Regions; Up-Regulation; Variant; Work; adipocyte biology; adipokines; adrenergic; basal insulin; cytokine; energy balance; feeding; human FRAP1 protein; human TNF protein; in vivo; interest; mRNA Stability; mTOR Signaling Pathway; messenger ribonucleoprotein; nucleocytoplasmic transport; nucleoprotein kinase; programs; protein expression; protein kinase R; research study; response

DESCRIPTION (provided by applicant): The adipocyte hormone leptin regulates energy balance, substrate metabolism, immunity, bone formation and reproduction. Our recent work demonstrates that leptin production is regulated at the translational level, and that elements within its 5' UTR stimulate and its 3' UTRs inhibit translation of a reporter gene. The major goal of this application is to identify the cis elements and trans acting factors that regulate leptin translation in response to variations in nutritional state. The objectives of this application are: 1) To identify RNA binding proteins (RBP) that interact with the leptin 5' and 3'UTRs, and define the cis elements involved; 2) To determine the effect of starvation/feeding in vivo and acute treatment with insulin and beta-adrenergic agonists in vitro on expression of RBPs and their interaction with leptin mRNA; 3) To assess the functional roles of specific RBPs in leptin mRNA translation using knockdown and overexpression studies; and 4) To delineate the signaling mechanisms that cause high basal and insulin-resistant leptin translation in obesity. The leptin 3'UTR includes AU-rich sequences and motifs for binding of RBPs that are known to stimulate (HuR) or inhibit (TIA-1, TIAR) the translation of specific mRNAs. We will therefore determine if leptin mRNA 'in vivo' associates with RBPs 'in vivo' by immunoprecipitating ribonucleoprotein complexes from cytosolic extracts of 3T3-L1 adipocytes and rat adipocytes. Pull-down assays using biotinylated leptin UTR probes will verify the interactions of RBPs of interest and point to the motifs involved. We will test the hypothesis that insulin in vitro or nutritional status in vivo (starvation, feeding) affects the binding of specific RBPs to leptin mRNA, and involves an alteration in their expression and/or nucleocytoplasmic shuttling. Finally, we will test the hypothesis that the chronic hyperinsulinemia associated with obesity increases leptin production at the translational level through the activation of stress / energy sensing pathways (e.g. mTOR, AMPK, and MAPK) and/or by altering the expression or binding of specific RBPs to leptin mRNA. Overall, the proposed studies will enhance understanding of leptin biology and broaden knowledge of how the adipocyte functions as an endocrine cell that orchestrates the production of numerous hormones and cytokines in response to nutritional cues.

描述（申请人提供）：脂肪细胞激素瘦素调节能量平衡、底物代谢、免疫、骨形成和繁殖。我们最近的工作表明，瘦素的产生在翻译水平上受到调节，并且其 5' UTR 内的元件刺激报告基因的翻译，而其 3' UTR 则抑制报告基因的翻译。该应用的主要目标是确定调节瘦素翻译以响应营养状态变化的顺式元件和反式作用因子。本申请的目标是： 1) 鉴定与瘦素 5' 和 3'UTR 相互作用的 RNA 结合蛋白 (RBP)，并定义所涉及的顺式元件； 2) 确定体内饥饿/喂养以及体外胰岛素和β-肾上腺素能激动剂急性治疗对RBP表达及其与瘦素mRNA相互作用的影响； 3) 使用敲低和过表达研究评估特定 RBP 在瘦素 mRNA 翻译中的功能作用； 4) 描述导致肥胖症基础高和胰岛素抵抗瘦素翻译的信号传导机制。瘦素 3'UTR 包含富含 AU 的序列和基序，用于结合已知可刺激 (HuR) 或抑制 (TIA-1、TIAR) 特定 mRNA 翻译的 RBP。因此，我们将通过免疫沉淀来自 3T3-L1 脂肪细胞和大鼠脂肪细胞胞浆提取物的核糖核蛋白复合物来确定瘦素 mRNA“体内”是否与 RBP“体内”相关。使用生物素化瘦素 UTR 探针的 Pull-down 测定将验证感兴趣的 RBP 的相互作用并指出所涉及的基序。我们将检验以下假设：体外胰岛素或体内营养状态（饥饿、进食）会影响特定 RBP 与瘦素 mRNA 的结合，并涉及其表达和/或核质穿梭的改变。最后，我们将检验以下假设：与肥胖相关的慢性高胰岛素血症通过激活应激/能量传感途径（例如 mTOR、AMPK 和 MAPK）和/或通过改变特定 RBP 的表达或与瘦素 mRNA 的结合，在翻译水平上增加瘦素的产生。总体而言，拟议的研究将增强对瘦素生物学的理解，并拓宽对脂肪细胞如何作为内分泌细胞发挥作用的知识，该内分泌细胞根据营养提示协调产生多种激素和细胞因子。