Transcriptional Regulation of SREBP-1c by Dietary Polyunsaturated Fatty Acids

膳食多不饱和脂肪酸对 SREBP-1c 的转录调节

• 批准号: 7615186
• 负责人: George E Howell
• 金额: $ 0.86万
• 依托单位: UNIVERSITY OF TENNESSEE HEALTH SCI CTR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-01-01 至 2009-02-12
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7615186
• 关键词: Acetyl-CoA Carboxylase; Animal Model; Animals; Binding; Binding Proteins; Biological Assay; CREB-binding protein; Dietary Supplementation; Dyslipidemias; EP300 gene; Elements; Elevation; Face; Fatty Acids; Fatty-acid synthase; Gene Targeting; Genes; Genetic Transcription; Hepatic; Hepatocyte; Hyperinsulinism; In Vitro; Insulin; Laboratories; Liver; Luciferases; Mediating; Menhaden oil; Molecular; Mutation; Non-Insulin-Dependent Diabetes Mellitus; Numbers; Obesity; Phosphorylation; Plasmids; Polymerase Chain Reaction; Polyunsaturated Fatty Acids; Production; Promoter Regions; Protein Overexpression; Proteins; Publishing; Rattus; Regulatory Element; Repression; SRE-1 binding protein; Small Interfering RNA; Sp1 Transcription Factor; Steroid Receptors; Sterols; Supplementation; Time; Trans-Activators; Transcription Coactivator; Transcriptional Activation; Transcriptional Regulation; Western Blotting; base; chromatin immunoprecipitation; combinatorial; feeding; gene repression; human CREBBP protein; in vivo; insight; lipid biosynthesis; novel; nuclear factor Y; promoter; receptor; transcription factor

DESCRIPTION (provided by applicant): Sterol regulatory element binding protein (SREBP-1c) is the primary transcription factor that governs expression of a host of lipogenic genes in the liver. In states of obesity and hyperinsulinemia, SREBP-1c production is increased due, in part, to elevated levels of transcription. First, an animal model of obesity and hyperinsulinemia, the JCR:LA-cp rat, will be employed to determine the effect of dietary supplementation with menhaden oil on the in vivo associations of known transcription factors and coactivators (LXRa, NF-Y, Sp-1, SREBP-1c, SRC-1, CBP/p300) and possible repressers (Kid-1 and NrOb2) with the promoter region of SREBP-1c and its target genes by chromatin immunoprecipitation assays. The effect of menhaden oil supplementation on abundance of these factors will be determined by real-time PCR and western blotting. In addition, novel PUFA regulatory mechanisms will be examined such as transcriptional repression by NrOb2, Kid-1, and phosphorylation of insulin-sensitive transcription factors/coactivators. Secondly, to determine the molecular mechanism(s) by which PUFA repress insulin stimulated SREBP-1c transcription, mutational analysis of transcription factor binding elements within the rat SREBP-1c promoter will be performed to dissect the elements which mediate PUFA effects. Rat SREBP-1c promoter driven luciferase assays in rat primary hepatocytes will be used to determine transcriptional activity. Gene knockdown by siRNA directed against target proteins will be used to confirm promoter mutation studies and plasmid mediated overexpression of candidate transactivators will be used to "rescue" transcriptional activity in the face of PUFAs. Taken together, the presently proposed studies should provide much needed insight into the molecular mechanism(s) through which dietary PUFAs can ameliorate elevations in SREBP-lc expression and the resulting dyslipidemia observed in obesity and hyperinsulinemia. Relevance: The current proposal seeks to determine the mechanism(s) by which long chain polyunsaturated fatty acids (PUFAs) decrease expression of SREBP-1c and lipogenesis in the obese state thereby ameliorating dyslipidemia associated with obesity and type II diabetes.

描述（由申请方提供）：固醇调节元件结合蛋白（SREBP-1c）是控制肝脏中脂肪生成基因宿主表达的主要转录因子。在肥胖和高胰岛素血症的状态下，SREBP-1c的产生增加，部分原因是转录水平升高。首先，将采用肥胖和高胰岛素血症的动物模型JCR：LA-cp大鼠，通过染色质免疫沉淀测定法，确定膳食补充鲱鱼油对已知转录因子和共激活因子（LXR α、NF-Y、Sp-1、SREBP-1c、SRC-1、CBP/p300）以及可能的阻遏物（Kid-1和NrOb 2）与SREBP-1c及其靶基因的启动子区域的体内关联的影响。鲱鱼油补充对这些因子丰度的影响将通过实时PCR和蛋白质印迹来确定。此外，新的PUFA调节机制将被检查，如转录抑制NrOb 2，Kid-1，和胰岛素敏感的转录因子/辅激活因子的磷酸化。其次，为了确定PUFA抑制胰岛素刺激的SREBP-1c转录的分子机制，将对大鼠SREBP-1c启动子内的转录因子结合元件进行突变分析，以剖析介导PUFA作用的元件。将使用大鼠原代肝细胞中的大鼠SREBP-1c启动子驱动的荧光素酶测定来确定转录活性。针对靶蛋白的siRNA基因敲低将用于确认启动子突变研究，而质粒介导的候选反式激活因子过表达将用于在PUFA面前“拯救”转录活性。综上所述，目前提出的研究应该提供对膳食PUFA可以改善SREBP-lc表达的升高以及在肥胖和高胰岛素血症中观察到的所得血脂异常的分子机制的急需的洞察。相关性：目前的提议旨在确定长链多不饱和脂肪酸（PUFA）在肥胖状态下降低SREBP-1c表达和脂肪生成从而改善与肥胖和II型糖尿病相关的血脂异常的机制。