Characterization of human IRS-1 Phosphorylation at S348, T495, and S1005

描述(申请人提供)：我们实验室的研究重点是人类骨骼肌中胰岛素抵抗的机制。具体地说，我们询问信号蛋白IRS-1的丝氨酸/苏氨酸磷酸化如何调节该蛋白的功能，这是肌肉中胰岛素作用的关键。利用灵敏的质谱学技术，我们鉴定了从人股外侧肌分离的IRS-1的22个丝氨酸/苏氨酸磷酸化位点。我们量化了胰岛素对其中15个残基的体内磷酸化的影响。在这15个位点中，4个残基Ser348、Thr446、Thr495和Ser1005的磷酸化在胰岛素治疗后降低。这项建议的目的是为朗莱博士提供支持，以确定在肥胖和2型糖尿病患者中，与瘦的健康对照组相比，人类IRS-1中胰岛素控制的模式和水平以及人类IRS-1中Ser348、Thr446、Thr495和Ser1005的磷酸化是否被解除调控。这项建议的第二个目的是确定负责它们磷酸化的激酶(S)，mTOR是主要的候选激酶。这些实验将提供一个概念性的框架，将我们实验室关于在活体中调节人类肌肉中这些位置的磷酸化的其他结果放在其中。因此，朗莱斯博士的实验将与实验室的更大使命交织在一起。胰岛素抵抗的分子机制的定义将提供有关2型糖尿病发病机制的知识，并有可能为治疗方式提供分子靶点。 与公共卫生相关：了解胰岛素如何在其靶组织(如骨骼肌)中无法正常发挥作用，将有助于深入了解个体如何发展胰岛素抵抗、2型糖尿病、肥胖和代谢综合征。由于这些疾病是一个日益严重的公共卫生问题，加起来约占公共卫生成本的40%，因此必须了解这些疾病的起源、预防和可能的治疗方法。

DESCRIPTION (provided by applicant): Research in our laboratory focuses on the mechanisms of insulin resistance in human skeletal muscle. Specifically, we ask how serine/threonine phosphorylation of the signaling protein IRS-1 modulates the function of this protein, which is a key to insulin action in muscle. Using sensitive mass spectrometry techniques, we identified 22 serine/threonine phosphorylation sites in IRS-1 isolated from human vastus lateralis muscle. We quantified the effect of insulin on the in vivo phosphorylation of 15 of these residues. Of these 15 sites, phosphorylation of 4 residues, Ser348, Thr446, Thr495, and Ser1005, was decreased upon insulin treatment. The purpose of this proposal is to provide support for Dr. Langlais to ascertain whether the insulin controlled pattern and level and of phosphorylation of Ser348, Thr446, Thr495, and Ser1005 in human IRS-1 is deregulated in obese and type 2 diabetic human subjects as compared to lean, healthy controls. The second aim of this proposal is to identify the kinase(s) responsible for their phosphorylation, with mTOR being the principal candidate kinase. These experiments will provide a conceptual framework in which to place into context other results from our laboratory regarding regulation of phosphorylation of these sites in human muscle in vivo. Therefore Dr. Langlais' experiments will interweave with the larger mission of the laboratory. Definition of the molecular mechanisms responsible for insulin resistance will provide knowledge regarding the pathogenesis of type 2 diabetes mellitus and potentially provide molecular targets for treatment modalities. PUBLIC HEALTH RELEVANCE: Understanding how insulin fails to function properly in its target tissues, such as skeletal muscle, will provide insight into how individuals develop insulin resistance, type 2 diabetes, obesity, and the metabolic syndrome. Since these diseases are a growing public health problem responsible together for about 40% of public health costs, knowledge of the origins prevention and potential treatments for these diseases are imperative.