ERK Kinase Regulation of Insulin Signaling and Inflammation: Honing the Antidiabetic Effects of PPAR-gamma Activation

 描述（由申请人提供）：噻唑烷二酮类（TZD）是强效抗糖尿病药物，其在治疗2型糖尿病中的应用受到不良副作用的限制。该提案的目标是对一种机制进行生物化学研究，该机制将TZD的积极代谢作用与其副作用分开，从而有可能设计出更好的一类药物。TZD的分子靶点PPAR γ是全身胰岛素敏感性、脂肪生成、炎症和能量稳态的关键调节因子。在脂肪组织中，在高脂饮食开始后不久，观察到PPAR γ在丝氨酸273（S273）处的磷酸化。 饮食喂养，并随着进行性肥胖而增加。这种磷酸化与PPAR γ靶基因的失调相关，例如胰岛素增敏激素脂联素的表达降低。ERK是负责磷酸化PPAR γ S273的主要激酶，MEK/ERK激酶通路的抑制剂阻断PPAR γ S273磷酸化。令人惊讶的是，MEK/ERK抑制剂在肥胖小鼠中具有有效的抗糖尿病作用，显示出显著改善的葡萄糖稳态。类似地，缺乏促进脂肪形成能力但仍然阻断S273磷酸化的高亲和力的PPAR γ γ配体保留了抗糖尿病作用。这两种阻断PPAR γ S273磷酸化的药理学干预均促进改善外周代谢稳态（如TZD），同时也更安全，因为它们不会引发与TZD相关的副作用。我们的假设是，肥胖和炎症中ERK介导的PPAR γ磷酸化通过靶向脂肪组织转录调节导致葡萄糖稳态受损。我们将使用一种新的转基因小鼠来检验这一假设，这种小鼠的S273（S273A）上的PPAR γ不能磷酸化。我们的初步数据表明，阻断这种磷酸化足以改善肥胖症患者的胰岛素敏感性。在本提案中，我们将利用三种方法来理解 PPAR γ磷酸化在肥胖发病机制中的作用。在目的1中，我们将研究PPAR γ S273磷酸化对葡萄糖稳态的影响以及PPAR γ S273A小鼠对PPAR γ配体的反应能力。在目标2中，我们将研究阻断免疫系统中S273磷酸化对脂肪组织炎症和胰岛素抵抗的相对贡献。在目的3中，我们将检测与野生型PPAR γ或磷酸化非依赖性S273A PPAR γ结合的mRNA转录调控元件和顺式调控元件的全基因组集合。了解肥胖和炎症如何通过ERK介导的磷酸化调节PPAR γ对于未来设计新的治疗分子将是重要的。

 DESCRIPTION (provided by applicant): The thiazolidinediones (TZDs) are powerful anti-diabetic drugs whose use in treating type 2 diabetes is limited by adverse side effects. The goal of this proposal is a biochemical investigation into a mechanism that separates the positive metabolic effects of TZDs from their side effects, making it possible to design a better class of agents. PPARγ, the molecular target of the TZDs, is a key regulator of systemic insulin sensitivity, adipogenesis, inflammation, and energy homeostasis. In adipose tissue, phosphorylation of PPARγ at serine 273 (S273) is observed shortly after the initiation of high fat diet feeding and increases with progressive obesity. This phosphorylation correlates with dysregulation of PPARγ target genes, such as decreased expression of the insulin-sensitizing hormone adiponectin. ERK is the primary kinase responsible for phosphorylating PPARγ S273, inhibitors of the MEK/ERK kinase pathway block PPARγ S273 phosphorylation. Surprisingly, MEK/ERK inhibitors had potent anti-diabetic effects in obese mice demonstrating markedly improved glucose homeostasis. Similarly, high-affinity ligands of PPARγwhich lack the capacity to promote adipogenesis but still block S273 phosphorylation retain anti-diabetic effects. These two pharmacological interventions blocking PPARγ S273 phosphorylation both promote improved peripheral metabolic homeostasis like the TZDs while also appearing safer as they do not trigger the side effects associated with TZDs. Our hypothesis is that ERK-mediated phosphorylation of PPARγ in obesity and inflammation causes altered impaired glucose homeostasis by targeting adipose tissue transcriptional regulation. We will test this hypothesis using a novel genetically modified mouse where PPARγ cannot be phosphorylated on S273 (S273A). Our preliminary data suggest that blocking this phosphorylation is sufficient to improve insulin sensitivity in obesity. In this proposal we will utilize three approaches to understand the contribution of PPARγ phosphorylation to the pathogenesis of obesity. In Aim 1, we will interrogate the effects of PPARγ S273 phosphorylation on glucose homeostasis and the ability of PPARγ S273A mice to respond to PPARγ ligands. In Aim 2, we will investigate the relative contribution of blocking S273 phosphorylation in the immune system to adipose tissue inflammation and insulin resistance. In Aim 3, we will examine the genome-wide set of mRNA transcripts regulated and cis-regulatory elements bound to either wild-type PPARγ or phosphorylation independent S273A PPARγ. Understanding how obesity and inflammation modulates PPARγ by ERK- mediated phosphorylation will be important for the future design of new therapeutic molecules.