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Proteomics analysis for the novel liver specific transcription factors and clarification of their control signals

新型肝脏特异性转录因子的蛋白质组学分析及其控制信号的澄清

基本信息

批准号：
17590941
负责人：
MIYAMURA Nobuhiro
金额：
$ 2.24万
依托单位：
KUMAMOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

项目摘要

Insulin is essential for maintaining glucose homeostasis. Insulin binds to the insulin receptor and activates its endogenous tyrosine kinase, which in turn phosphorylates insulin receptor substrates (IRSs), and transmits various downstream signals. The liver is one of the major target organs of insulin in which the expression of the insulin receptor is abundant. It is suggested that the expression of insulin receptor is regulated by tissue specific mechanism. However, the detail is still uncertain. On the other hand, AMP-activated protein kinase (AMPK) has been known to be important in the regulation of glucose and lipid metabolism in liver. The aim of this study is to investigate the effect of AMPK on the insulin receptor, which has a key role in the insulin action, in liver cells that is the major target organ of both AMPK and insulin. We investigated the effect of 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR), which is an activator of AMPK, on the expression of insulin … More receptor in a human hepatoma cell line, HepG2 cells. AICAR treatment for 48 hours significantly decreased the expression of insulin receptor protein in a dose-dependent manner in HepG2 cells. On the other hand, the inhibitory effect of AICAR was not observed in either 3T3-L1 adipocytes or CHO cells. The expression of insulin receptor mRNA was also significantly decreased with AICAR treatment in a dose-dependent manner. In addition, transcriptional activity of the insulin receptor gene promoter was also down-regulated with AICAR treatment. The inhibitors of AICAR blocked the effects of AICAR on the down-regulation of insulin receptor protein, mRNA and promoter activity. According to the investigation with a deletion mutant of the insulin receptor gene promoter, it was suggested that cis-elements responsible for the AICAR-induced down-regulation existed within 0.6 kb upstream from the ATG codon in the insulin receptor gene. We have found five consensus sequences of insulin response element (IRE-1〜5) in the human insulin receptor gene promoter. A transcription factor Foxol was suggested to bind to IRE-4 and IRE-5, which exist within 0.6 kb of AICAR response region, and the binding to each IREs was decreased with AICAR treatment. This study demonstrated for the first time that AMPK activation reduced the expression of insulin receptor, at least in part, by a down-regulation of the gene transcription, and that this effect may be specific in liver cells. In addition, it was suggested that Foxol was involved in the transcriptional regulation of insulin receptor gene in liver cells. Less

胰岛素是维持葡萄糖稳态所必需的。胰岛素与胰岛素受体结合并激活其内源性酪氨酸激酶，酪氨酸激酶进而磷酸化胰岛素受体底物（IRS），并传递各种下游信号。肝脏是胰岛素的主要靶器官之一，其中胰岛素受体表达丰富。提示胰岛素受体的表达受组织特异性机制的调控。不过，具体细节还不确定。另一方面，AMP激活的蛋白激酶（AMPK）在肝脏糖脂代谢的调节中起重要作用。本研究的目的是研究AMPK对胰岛素受体的影响，胰岛素受体在胰岛素作用中起关键作用，肝细胞是AMPK和胰岛素的主要靶器官。我们研究了AMPK激活剂5-氨基咪唑-4-甲酰胺-1-β-D-呋喃核糖苷（AICAR）对胰岛素表达的影响 ...更多信息受体在人肝癌细胞系HepG 2细胞中的表达。AICAR处理48 h后，HepG 2细胞胰岛素受体蛋白表达呈剂量依赖性下降。另一方面，AICAR在3 T3-L1脂肪细胞或CHO细胞中均未观察到抑制作用。AICAR处理组胰岛素受体mRNA表达量明显降低，且呈剂量依赖性。此外，AICAR处理也下调了胰岛素受体基因启动子的转录活性。AICAR抑制剂可阻断AICAR对胰岛素受体蛋白、mRNA和启动子活性的下调作用。根据对胰岛素受体基因启动子缺失突变体的研究，提示在胰岛素受体基因ATG密码子上游0.6kb内存在AICAR诱导下调的顺式元件。我们在人胰岛素受体基因启动子中发现了5个胰岛素反应元件（IRE-1 β 5）的共有序列。在AICAR反应区的0.6kb范围内，有转录因子Foxol与IRE-4和IRE-5结合，AICAR处理后，Foxol与IRE-4和IRE-5的结合减弱。这项研究首次证明AMPK激活至少部分通过下调基因转录降低胰岛素受体的表达，并且这种作用可能在肝细胞中具有特异性。此外，Foxol可能参与了肝细胞胰岛素受体基因的转录调控。少