HORMONAL AND NUTRITIONAL CONTROL OF ENZYME BIOSYNTHESIS

酶生物合成的激素和营养控制

• 批准号: 3234594
• 负责人: Hei Sook Sul
• 金额: $ 5.48万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-03-01 至 1994-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3234594
• 关键词: 6 phosphofructokinase; DNA footprinting; adipocytes; affinity chromatography; chemical models; enzyme biosynthesis; enzyme induction /repression; fatty acid synthase; gel mobility shift assay; gene expression; gene mutation; genetically modified animals; hormone regulation /control mechanism; insulin; laboratory mouse; molecular cloning; nucleic acid sequence; site directed mutagenesis; tissue /cell culture; transcription factor; transfection

Glucose utilization via glycolysis and lipogenesis must be regulated to meet the changing nutritional state of the animals; glycolysis is low and fatty acid synthesis is virtually absent during fasting, and fatty acids and triacylglycerol synthesis for energy storage increase dramatically during feeding. Secretion of insulin, a major anabolic polypeptide hormone, is increased during feeding and is involved in this regulation. In addition, glycolysis and fatty acid synthesis is impaired in diabetic animals and insulin restores the rates to normal levels. Since diabetes is among the most common and serious metabolic diseases, understanding mechanisms of insulin action is of utmost importance. The goal of this research is to determine the molecular mechanisms by which insulin increases transcription of two key enzymes of energy metabolism. Fatty acid synthase (FAS) plays central role in de novo lipogenesis by catalyzing all the reaction in the conversion of acetyl CoA and malonyl CoA to palmitate. Phosphofructokinase-1 (PFK) catalyzes the committed step in glycolysis. We have cloned cDNA and genomic clones for FAS, liver PFK and other insulin-inducible mRNAs and have shown transcriptional stimulation of these genes by insulin. The 5'-flanking regions of FAS and PFK genes contained sequences which confer insulin activation to linked reporter genes when transfected into 3T3-Ll adipocytes. A common mechanism may be involved in the insulin induction of these enzymes. Our goal is to define cis-acting elements and trans- acting factors for the transcriptional activation of these genes by insulin. Using in vitro mutagenesis, essential sequences for insulin regulation will be defined by transfection into adipocytes and by utilizing transgenic mice. The nuclear factor(s) which interact with these sequences will be characterized by mobility shift and DNase footprinting assays. The factor(s) will then be purified and/or cloned to start studying at the molecular level how insulin stimulates transcription of these genes. This will lead us in the future to elucidate a signal transduction pathway of insulin.

必须通过糖酵解和脂肪形成来调节葡萄糖的利用 满足不断变化的动物营养状态；糖酵解低，并且 禁食过程中实际上不存在脂肪酸合成，而脂肪酸则是脂肪酸的 和三酰基甘油合成用于能量储存的合成急剧增加 在喂食期间。 胰岛素的分泌，一种主要的合成代谢多肽 激素在进食过程中增加，并参与该调节。 另外，糖尿病患者糖酵解和脂肪酸合成受损 动物和胰岛素将速率恢复到正常水平。 由于糖尿病 是最常见和最严重的代谢疾病之一，理解 胰岛素作用的机制至关重要。 这项研究的目的是通过 胰岛素增加了两个关键能量酶的转录 代谢。 脂肪酸合酶（FAS）在从头上起着核心作用 通过催化乙酰基转化中的所有反应，脂肪生成 COA和Malonyl COA到棕榈酸酯。 磷酸果果酶1（PFK）催化 致力于糖酵解的步骤。 我们有克隆的cDNA和基因组克隆 对于FAS，肝脏PFK和其他胰岛素诱导的mRNA，已显示 通过胰岛素对这些基因的转录刺激。 5'FARKING FA和PFK基因的区域包含赋予胰岛素的序列 转染到3T3-ll时激活链接的报告基因 脂肪细胞。 胰岛素诱导可能涉及常见机制 这些酶。 我们的目标是定义顺式作用元素和反式元素 通过转录激活这些基因的作用因素 胰岛素。 使用体外诱变，胰岛素必需序列 调节将通过转染到脂肪细胞和 利用转基因小鼠。 与之相互作用的核因子 这些序列将以移动性转移和DNase为特征 足迹测定法。 然后将纯化和/或克隆该因子 开始在分子水平学习胰岛素如何刺激 这些基因的转录。 这将使我们将来 阐明胰岛素的信号转导途径。