The role of serine phosphorylation of IRS-2 in hepatic insulin resistance

项目摘要 本修订提案（DK 081513）侧重于研究IRS-2的丝氨酸/苏氨酸磷酸化， ERK、mTOR和p70 S6 K介导初始肝胰岛素抵抗胰岛素在肝脏中的作用 在维持葡萄糖稳态中的重要作用。肝脏胰岛素信号传导缺陷有助于系统性 胰岛素抵抗和2型糖尿病的慢性高血糖。在肝脏中，IRS-2是 胰岛素受体酪氨酸激酶的生理底物;因此，IRS-2具有足够的功能性 对肝脏胰岛素信号传导至关重要。IRS-2含有许多潜在的丝氨酸/苏氨酸磷酸化 这些位点可能被多种丝氨酸/苏氨酸激酶靶向。IRS-1的丝氨酸磷酸化 已经显示出减弱胰岛素信号传导;然而，对IRS-2知之甚少。我们的初步研究表明 证实了许多丝氨酸/苏氨酸激酶的活化以及多种 胰岛素抵抗动物肝脏IRS-2中丝氨酸/苏氨酸残基。在测试过的少数已知的 IRS-1、ERK和mTOR/p70 S6 K激酶是唯一与肝胰岛素抵抗相关的激酶， 表明这些丝氨酸激酶在抗性肝脏中被选择性激活。在本提案中，我们 假设IRS-2通过ERK、mTOR和/或p70 S6 K丝氨酸/苏氨酸磷酸化形成了一个 肝脏中的初始胰岛素抵抗信号并引起肝脏胰岛素抵抗，导致外周胰岛素 阻力该应用程序旨在在肝脏和分离的原代肝细胞中检验这一假设。 这些激酶的活化及其对IRS-2的磷酸化在细胞凋亡过程中的动态变化， 将监测胰岛素抗性的发展。关键的丝氨酸/苏氨酸激酶将被鉴定， 与胰岛素信号通路的相互作用-特别是IRS-2-将使用腺病毒进行研究。 和/或siRNA技术。独特的策略也将用于鉴定未知的丝氨酸/苏氨酸 IRS-2和IRS-2激酶中的磷酸化位点。本提案的目的是定义初始胰岛素 抗信号（激酶活化和IRS-2磷酸化）。这次调查的结果 应该提供一个更清晰的，综合的理解胰岛素敏感性的分子决定因素， 肝脏以及整个机体。

Project Summary This revised proposal (DK081513) focuses on the investigation of serine/threonine phosphorylation of IRS-2 by ERK, mTOR and p70S6K in mediating initial hepatic insulin resistance. Insulin action in the liver plays an important role in maintaining glucose homeostasis. Defects in hepatic insulin signaling contribute to systematic insulin resistance and the chronic hyperglycemia of type 2 diabetes. In the liver, IRS-2 is the primary physiological substrate for the insulin receptor tyrosine kinase; as such, sufficient functionality of IRS-2 is essential for proper hepatic insulin signaling. IRS-2 contains many potential serine/threonine phosphorylation sites which are potentially targeted by diverse serine/threonine kinases. Serine phosphorylation of IRS-1 has been shown to attenuate insulin signaling; however, little is known about IRS-2. Our preliminary studies have confirmed activation of numerous serine/threonine kinases as well as phosphorylation of multiple serine/threonine residues in IRS-2 from the liver of insulin-resistant animals. Among tested handful known kinases for IRS-1, ERK and mTOR/p70S6K were the only kinases that correlated with hepatic insulin resistance, suggesting that these serine kinases are selectively activated in resistant liver. In this proposal, we hypothesize that serine/threonine phosphorylation of IRS-2 by ERK, mTOR and/or p70S6K forms an initial insulin resistant signal in liver and cause hepatic insulin resistance, leading to peripheral insulin resistance. This application is designed to test this hypothesis in liver and in isolated primary hepatocytes. Dynamic changes in the activation of these kinases and their phosphorylation on IRS-2 during the development of insulin resistance will be monitored. Key serine/threonine kinase will be identified and its interactions with the insulin signaling pathway -and IRS-2 in particular- will be investigated using adenoviral and/or siRNA techniques. Unique strategies will also be applied to identify unknown serine/threonine phosphorylation sites in IRS-2 and IRS-2 kinases. The objective of this proposal is to define the initial insulin resistant signal (activation of kinase and phosphorylation of IRS-2) in liver. The results from this investigation should provide a clearer, integrative understanding of the molecular determinants of insulin sensitivity in the liver as well as the whole organism.