Department of Internal Medicine II, Center for Molecular Medicine Cologne (CMMC), University of Cologne, Kerpener Strasse 62, 50937 Cologne, Germany.
J Mol Endocrinol. 2010 Feb;44(2):99-113. doi: 10.1677/JME-08-0182. Epub 2009 Sep 15.
Recent studies have discovered changes in the insulin-/IGF1 signaling affecting glucose metabolism and the molecular pathogenesis of human hepatocellular cancer. Insulin/IGF1 receptor mediates its intracellular effects by recruitment of one out of the four different insulin receptor substrates (IRS). To investigate mechanisms of IRS2 expression, we analyzed transcriptional regulation of IRS2 in human HepG2 cells. We identified a region 688 bp upstream of the translation start codon responsible for approximately 90% of basal human IRS2 promoter activity in HepG2 cells, and confirmed binding of specificity protein 1 (also called Sp1 transcription factor, SP1) and nuclear factor 1 (NFI) in this region. Mutation of both SP1 and NFI binding sites or inhibition of extracellular signal regulated kinase (ERK) suppressed IRS2 promoter activity almost completely, revealing a major role of MAP kinases (MAPK) for IRS2 transcription. Activating this cascade with oxidative stress increased IRS2 promoter activity and endogenous IRS2 expression substantially. IRS2 promoter activity rose even more after additional inhibition of p38MAPK indicating an inhibitory effect of p38MAPK on ERK mediated IRS2 transcription. Activation of the MAPK pathway using interleukin 1, beta (IL1B) increased IRS2 promoter activity similar to oxidative stress. In contrast IL1B decreases and inhibition of the MAPK pathway increases IRS1 promoter activity revealing opposed effects of IL1B and ERK on the expression of different IRS proteins. In conclusion we discovered a specific region (-688 to -611 bp) in the IRS2 promoter essential for basal promoter activity and oxidative stress induced transcription depending on ERK activation and SP1 and NFI binding in human hepatocytes.
最近的研究发现，胰岛素/IGF1 信号通路的变化会影响葡萄糖代谢和人类肝细胞癌的分子发病机制。胰岛素/IGF1 受体通过募集四种不同的胰岛素受体底物（IRS）之一来介导其细胞内效应。为了研究 IRS2 表达的机制，我们分析了人 HepG2 细胞中 IRS2 的转录调控。我们确定了翻译起始密码子上游 688 个碱基的区域，该区域负责 HepG2 细胞中基础人 IRS2 启动子活性的约 90%，并在该区域证实了特异性蛋白 1（也称为 Sp1 转录因子，SP1）和核因子 1（NFI）的结合。SP1 和 NFI 结合位点的突变或细胞外信号调节激酶（ERK）的抑制几乎完全抑制了 IRS2 启动子活性，揭示了 MAP 激酶（MAPK）在 IRS2 转录中的主要作用。用氧化应激激活该级联反应会大大增加 IRS2 启动子活性和内源性 IRS2 表达。在另外抑制 p38MAPK 后，IRS2 启动子活性甚至进一步升高，表明 p38MAPK 对 ERK 介导的 IRS2 转录具有抑制作用。使用白细胞介素 1β（IL1B）激活 MAPK 通路会增加 IRS2 启动子活性，类似于氧化应激的作用。相反，IL1B 降低并且 MAPK 通路的抑制增加 IRS1 启动子活性，表明 IL1B 和 ERK 对不同 IRS 蛋白的表达具有相反的作用。总之，我们在人肝细胞中发现了 IRS2 启动子中的一个特定区域（-688 至-611 bp），该区域对于基础启动子活性和氧化应激诱导的转录是必需的，这取决于 ERK 的激活以及 SP1 和 NFI 的结合。
