Mechanism of Gdf3 action to limit insulin sensitivity in obesity

项目摘要 噻唑烷二酮类（TZDs）是一种有效的抗糖尿病药物，但在2型糖尿病治疗中的应用受到限制 不良副作用。这项提议的目的是对这种机制进行生物化学研究， 将TZD降低血糖的积极代谢作用与其充分表征的消极作用分开 方面的影响. TZDs的分子靶点PPARγ通过促进胰岛素敏感性的形成来调节全身胰岛素敏感性。 新的脂肪细胞然而，我们发现TZDs对PPARγ有第二种生化功能，即阻断PPARγ的表达。 丝氨酸273（pS273）的磷酸化。我们发现，在丝氨酸273磷酸化的过氧化物酶体增殖物激活受体γ促进胰岛素分泌， 而不影响脂肪形成。逆转这种磷酸化与药理学或遗传 抑制足以促进胰岛素敏感性并增加对冷的产热反应。我们建议 PPARγ在丝氨酸273处的磷酸化促进胰岛素抵抗的主要机制之一是 通过增加生长分化因子3（Gdf 3）的表达。Gdf 3是TGF-β 1中的分泌蛋白。 BMP/BMP超家族和BMP信号传导的负调节因子。在脂肪组织中，BMP蛋白有助于 胰岛素敏感性和产热。因此，我们认为肥胖患者Gdf 3水平升高是导致肥胖的原因。 PPARγ S273磷酸化介导的胰岛素抵抗。事实上，我们表明，异位表达的 gdf 3足以引起瘦小鼠的胰岛素抵抗。在这个提议中，我们研究了Gdf 3的贡献， 肥胖和胰岛素抵抗的发病机制。我们预测，阻断Gdf 3水平或活性将恢复 全身胰岛素敏感性和促进产热。我们将询问Gdf 3对葡萄糖的作用 使用急性功能获得和功能丧失两种模型来研究体内平衡和肥胖。我们将利用创新 新的无线连续葡萄糖监测技术与间接量热法同时进行，以实现高 胰岛素抵抗、能量平衡和循环营养可用性的精确测量。我们将 确定Gdf 3是否是连接PPARγ S273磷酸化和胰岛素抵抗的机制。我们将 还确定Gdf 3是否需要BMP信号通过SMAD 1/5/8蛋白。最后，我们将执行一个 使用全基因组CRISPR/Cas9对Gdf 3信号传导所需的途径进行无偏见的研究 淘汰赛这一假设提出了一种新的治疗方式，利用特定的有益的 这是TZD治疗的一个方面，与TZD相关的副作用无关。该提案的结果将 明确确定Gdf 3是否是促进代谢健康的合适靶点。

Project Summary 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 the mechanism that separates the positive metabolic effects of TZDs to lower glucose from their well-characterized negative side effects. PPARγ, a molecular target of the TZDs, regulates systemic insulin sensitivity by promoting formation of new adipocytes. However, we show that the TZDs have a second biochemical function on PPARγ, to block phosphorylation of serine 273 (pS273). We find that phosphorylation of PPARγ at serine 273 promotes insulin resistance without affecting adipogenesis. Reversing this phosphorylation with pharmacological or genetic inhibition is sufficient to promote insulin sensitivity and increase thermogenic responses to cold. We propose that one of the main mechanisms by which phosphorylation of PPARγ at serine 273 promotes insulin resistance is through increased expression of growth differentiation factor 3 (Gdf3). Gdf3 is a secreted protein in the TGF- /BMP superfamily and a negative regulator of BMP signaling. In adipose tissue, BMP proteins contribute to insulin sensitivity and thermogenesis. We therefore propose that elevated levels of Gdf3 in obesity are the cause of insulin resistance mediated by PPARγ S273 phosphorylation. Indeed, we show that ectopic expression of Gdf3 is sufficient to cause insulin resistance in lean mice. In this proposal we examine the contribution of Gdf3 to the pathogenesis of obesity and insulin resistance. We predict that blocking Gdf3 levels or activity will restore whole-body insulin sensitivity and promote thermogenesis. We will interrogate the role of Gdf3 on glucose homeostasis and obesity using both acute gain-of-function and loss-of-function models. We will utilize innovative new wireless continuous glucose monitoring technology concurrently with indirect calorimetry to achieve high- precision measurements of insulin resistance, energy balance, and circulating nutrient availability. We will determine whether Gdf3 is the mechanism linking PPARγ S273 phosphorylation and insulin resistance. We will also determine whether Gdf3 requires BMP signaling through SMAD1/5/8 proteins. Finally, we will perform an unbiased investigation into the pathways required for Gdf3 signaling using a genome-wide CRISPR/Cas9 knockout screen. This hypothesis suggests a new therapeutic modality which harnesses a specific beneficial aspect of TZD treatment which is independent of TZD-associated side effects. The results from this proposal will definitively determine if Gdf3 is a suitable target for promoting metabolic health.