Role of DUSPs in adipocytes in response to inflammatory stress.

描述（由申请人提供）：2006年至2015年间，全球肥胖率预计将从16亿上升到23亿，而同期发达国家因糖尿病导致的死亡人数将增加80%以上。我们的长期目标是描述肥胖症发病与糖尿病发病率上升之间的分子和细胞机制。越来越多的证据表明，c-Jun N末端激酶（JNK）信号在炎症应激耦合肥胖到胰岛素抵抗（IR）过程中是一个关键的中介，但很少有研究探讨使这一途径失活性的机制，从而可能改善胰岛素敏感性。最近的证据表明，在炎症应激（如TNF）期间，双特异性磷酸酶（DUSPs）在JNK失活中的作用。我们的数据表明，在需要RNA合成的机制下，JNK活性在持续刺激下是短暂的，同时DUSP基因表达在对TNF的反应中增加。因此，我们提出了一个中心假设，即DUSPs以脂肪细胞表型特异性的方式调节局部JNK激活和生物学功能，以响应TNFalpha。了解DUSP对脂肪细胞中JNK的调节将有助于开发治疗肥胖和糖尿病的替代靶点。我们计划通过以下两个具体目标来检验我们的中心假设：目标1。研究DUSPs对脂肪细胞中JNK信号传导的机制作用。我们将通过以下方式来验证我们的中心假设：1)确定哪些dusp是诱导的，2)它们在哪里被区分开，以及3)它们以脂肪细胞表型特异性的方式在响应TNFalpha的局部生物学结果中发挥什么作用。目标2。阐明JNK活性在DUSP表达中的作用。我们将通过确定：1)哪些DUSP是JNK活性的下游靶点，2)JNK下游的哪个转录因子调节DUSP基因表达，以及3)JNK如何以表型特异性的方式调节DUSP1蛋白的稳定性来验证我们的假设。利用RNAi、异位表达、免疫印迹分析、qRT-PCR和葡萄糖摄取，我们将能够检查DUSPs对局部JNK活性和功能（如胰岛素抵抗）的作用，以及JNK及其下游转录因子对DUSPs的反馈调节。虽然在肥胖诱导的胰岛素抵抗和最终糖尿病过程中有几种途径传递炎症信号，但JNK途径已成为这些代谢性疾病的关键介质。本研究将建立新的调控JNK信号的治疗靶点，并为肥胖诱导糖尿病的治疗提供替代策略。

DESCRIPTION (provided by applicant): Globally, obesity rates are expected to rise from 1.6 billion to 2.3 billion between 2006 and 2015, while deaths due to diabetes will increase by over 80% in developed nations during the same time. Our long-term goal is to delineate the molecular and cellular the mechanisms that couple the onset of obesity with the rising incidence of diabetes. Mounting evidence established c-Jun N terminal kinase (JNK) signaling as a key mediator during infiammatory stress coupling obesity to insulin resistance (IR), yet few studies have examined mechanisms that deactivate this pathway, potentially improving insulin sensitivity. Recent evidence demonstrates a role for dual specificity phosphatases (DUSPs) in the deactivation of JNK during inflammatory stress (e.g. TNF). Our data demonstrate that JNK activity is transient in the presence of continuing stimuli via mechanisms that require RNA synthesis, with concomitant increases in DUSP gene expression in response to TNF. Therefore, we developed the central hypothesis that DUSPs regulate localized JNK activation and biological function in an adipocyte phenotype-specific manner in response to TNFalpha. Understanding DUSP regulation of JNK in adipocytes will allow for the development of alternative targets in the treatment of obesity and diabetes. We plan to test our central hypothesis by pursuing the following two specific aims: Aim 1. Examine the mechanistic role of DUSPs on JNK signaling in adipocytes. We will test our central hypothesis by: 1) establishing which DUSPs are induced, 2) where they are compartmentalized, and 3) what role they play on localized biological outcome in response to TNFalpha in an adipocyte phenotype-specific manner. Aim 2. Elucidate the role of JNK activity on DUSP expression. We will test our hypothesis by determining: 1) which DUSPs are downstream targets of JNK activity, 2) which transcription factor downstream of JNK regulates DUSP gene expression, and 3) how JNK regulates DUSP1 protein stability in a phenotype-specific manner. Using RNAi, ectopic expression, immunoblot analysis, qRT-PCR, and glucose uptake we will be able to examine the role of DUSPs on localized JNK activity and Function (e.g. insulin resistance), as well as feedback regulation ofthe DUSPs by JNK and its downstream transcription factors. While several pathways relay inflammatory signals during obesity-induced insulin resistance and ultimately diabetes, the JNK pathway has emerged as a critical mediator in these metabolic diseases. This study will establish new therapeutic targets that regulate JNK signaling, and provide alternative strategies for the treatment of obesity-induced diabetes.