Proinsulin ER export and beta cell ER homeostasis in health and diabetes

 描述(申请人提供)：到目前为止，在全基因组相关性研究中发现的大多数糖尿病相关基因都富含在胰腺β细胞中。胰岛β细胞衰竭在2型糖尿病(T2D)的发生和发展中起着核心作用。内质网是胰岛素原大量产生和加工的中心细胞器。内质网稳态对正常的β细胞功能至关重要，并通过蛋白质合成、折叠、输出和降解之间的微妙平衡来维持。相反，内质网稳态的破坏，由许多遗传和环境因素引起的糖尿病，诱导内质网应激并导致T2D中的β细胞死亡。与我们在β细胞内质网中蛋白质合成、折叠和降解方面的知识相比，内质网输出在胰岛素原生物生成和内质网稳态中的作用还知之甚少。这项提议解决了这一知识鸿沟。我们已经获得了强有力的初步结果，证明胰岛素原的内质网退出需要COPII(外壳蛋白复合体II)包被的小泡，而缺陷的COPII依赖的ER出口强烈地诱导ER应激。除了我们的结果，最近开始有证据表明，内质网高尔基体转运减少是导致脂肪毒性诱导的内质网应激的原因。在这些新发现的基础上，我们的目标是确定葡萄糖和脂肪毒性如何影响β细胞中蛋白质货物的内质网输出，特别是胰岛素原，以及内质网输出缺陷如何反过来促进内质网应激和β细胞功能障碍。超过一千种蛋白质，被称为蛋白质平衡网络，已经被提出维持内质网的动态平衡。然而，到目前为止，只有很少的ER蛋白被研究过，因为它们参与了内质网应激或β细胞中胰岛素原的生物生成。需要进行系统的蛋白质组学分析，以全面了解不同的糖尿病致病条件如何在T2D开始时扰乱β细胞ER的稳态。在这个提案中，我们将使用系统生物学的方法来检验中心假设，即糖脂毒性等致糖尿病因素通过扰乱ER蛋白的稳态和COPII依赖的ER输出来扰乱β细胞的ER功能。该提案有两个具体目标：1)确定COPII依赖的内质网出口在健康和糖尿病中是如何受到调节的；2)确定内质网伴侣和折叠酶在糖尿病中是如何改变的，以及这些变化如何影响β细胞的内质网出口。这项研究将使用最先进的定量蛋白质组学和其他系统生物学方法来表征糖尿病引起的条件不同地改变内质网稳态的分子机制。重点将放在内质网出口上，这是β细胞内质网稳态的一个未被研究的组成部分。

 DESCRIPTION (provided by applicant): Up to date, most of the diabetes-associated genes identified in genome-wide association studies are enriched in the pancreatic beta cells. Beta cell failure plays a central role in the development and progression of type 2 diabetes (T2D). The endoplasmic reticulum (ER) is a central organelle for the massive production and processing of proinsulin. ER homeostasis is critical for normal beta cell function and is maintained by the delicate balance between protein synthesis, folding, export and degradation. In contrast, the disruption of ER homeostasis, by many genetic and environmental diabetes-causing factors, induces ER stress and causes beta cell death in T2D. Compared to our knowledge in protein synthesis, folding and degradation in beta cell ER, the role of ER export in proinsulin biogenesis and ER homeostasis is much less understood. This proposal addresses this knowledge gap. We have obtained strong preliminary results demonstrating that ER exit of proinsulin requires COPII (coat protein complex II) coated vesicles and defective COPII dependent ER export strongly induces ER stress. In addition to our results, evidence has recently begun to emerge that reduced ER-Golgi transport is causative to lipotoxicity-induced ER stress in beta cells. On the basis of these novel findings, we aim to determine how glucose- and lipid-toxicity affect the ER export of protein cargo, particularly proinsulin, in beta cells and how defective ER export, in turn, contributes to ER stress and beta cell dysfunction. Over a thousand proteins, referred to as the proteostasis network, have been proposed to maintain ER homeostasis. However, to date, only a very small number of ER proteins have been investigated for their involvement in ER stress or proinsulin biogenesis in beta cells. A systematic proteomics analysis is needed to comprehensively understand how different diabetes-causing conditions perturb beta cell ER homeostasis at the onset of T2D. In this proposal, using systems biology approaches, we will test the central hypothesis that diabetogenic factors such as glucolipotoxicity disrupt beta cell ER function through perturbing ER protein homeostasis and COPII dependent ER export. There are two specific aims in this proposal: 1) Determine how COPII dependent ER export is regulated in health and diabetes; 2) Determine how ER chaperones and folding enzymes are altered in diabetes and how these changes affect beta cell ER export. This study will use state-of-the-art quantitative proteomics and other systems biology approaches to characterize the molecular mechanisms by which diabetes-causing conditions differentially alter ER homeostasis. The emphasis will be on the ER export, an under-investigated component of beta cell ER homeostasis.