Modifiers of Proinsulin Influence T2D Susceptibility

项目总结 2型糖尿病(T2D)是由于β细胞不能产生足够的胰岛素来维持正常血糖而引起的。 作为遗传/环境因素的结果，胰岛素抵抗的形成迫使贝塔细胞 增加胰岛素的产生。尽管贝塔细胞有一定的能力来补偿需求，但通过 在世界上约6亿肥胖患者中，约有三分之一的人会发展为糖尿病。 导致β细胞衰竭的因素尚不清楚。由于这种疾病的多基因性质，很可能 许多基因改变了β细胞的功能，任何单一基因的多态都不会被检测到，因为 他们做出了很小的贡献。我们的基本假设是，多个基因影响着 胰岛素原在内质网(ER)中折叠，并改变T2D的进程。值得注意的是，我们的 初步研究表明，在糖尿病之前，高脂肪饮食就足以导致胰岛素原错误折叠。 C57BL/6小鼠的发育。此外，我们还发现了可加重胰岛素原的基因修饰因子。 错误折叠和β细胞衰竭。我们假设，T2D中β细胞衰竭的根本原因是 内质网水平的崩溃，未能有效折叠过量的胰岛素原并导致 对下游加工和分泌的影响。为了检验我们的假设，我们成立了一个团队 一批杰出的研究人员共同努力，通过使用 最新的蛋白质组学、生物化学、细胞生物学、小鼠遗传学和生物信息学。在预赛中 我们开发了区分特定二硫键缺陷和其他错误折叠的方法的研究 胰岛素原的构象，产生了所有必要的小鼠菌株，并验证了蛋白质组质量 人胰岛胰岛素原相互作用的光谱分析方法。我们也展示了它的潜力 小分子以改善挑战的胰岛中胰岛素原的产生。我们希望我们的新方法将 识别胰岛素原折叠途径中的明显缺陷，这些缺陷代表了导致β细胞的最早变化 在小鼠模型和人类中都会死亡。我们R24拨款的三个目标集中在定义胰岛素原如何 当胰岛受到挑战时，折叠模式会发生变化，并确定蛋白质与胰岛素原的相互作用可能 预测胰岛素原通过分泌途径运输的效率，影响胰岛健康。目标1将 通过测量正常胰岛素原折叠过程中的中间产物来量化胰岛素原的折叠和未折叠状态 以及来自特征明确的小鼠模型的患病胰岛。目标2将定义与之相互作用的蛋白质 胰岛素原在从正常的、肥胖的非糖尿病人到人类供者的T2D胰岛过程中的变化。目标3 将阐明什么干预和伴侣功能可以保存生产性的胰岛素原折叠和 恢复有效的胰岛素原“蛋白质平衡”网络。总而言之，我们提议的研究可能会确定小说 T2D治疗干预的生物标志物和途径，因此对T2D的 NIDDK的使命。

PROJECT SUMMARY Type 2 diabetes (T2D) is caused by a failure of beta cells to produce sufficient insulin to maintain euglycemia. As a consequence of genetic/environmental factors, insulin resistance develops that pressures beta cells to increase insulin production. Although beta cells have some capacity to compensate for the demand, by approximately one-third of ~600 million individuals with obesity in the world develop go on to develop diabetes. The factors that lead to beta cell failure are unknown. Due to the polygenic nature of the disease, it is likely many genes modify beta cell function, and polymorphisms in any single gene would not be detected because they present minor contributions. Our underlying hypothesis is that multiple genes impact the efficiency of proinsulin folding in the endoplasmic reticulum (ER) and modify the progression of T2D. Significantly, our preliminary studies show that a high fat diet is sufficient to cause proinsulin misfolding well before diabetes development in C57BL/6 mice. In addition, we have identified genetic modifiers that exacerbate proinsulin misfolding and beta cell failure. We hypothesize that the fundamental cause of beta cell failure in T2D is a breakdown at the level of the ER with failure to efficiently fold excessive amounts of proinsulin and resulting consequences on downstream processing and secretion. To test our hypothesis, we have established a team of outstanding investigators to work together to identify critical proteins that modify proinsulin folding using state-of-the-art proteomics, biochemistry, cell biology, murine genetics and bioinformatics. In preliminary studies we developed methods to differentiate between specific disulfide bond defects and other misfolded conformations of proinsulin, generated all of the necessary murine strains and validated the proteomic mass spectrometry approach for proinsulin interactions using human islets. We have also demonstrated the potential of small molecules to improve proinsulin production in challenged islets. We expect our novel approach will identify distinct defects in the proinsulin folding pathway that represent the earliest changes leading to beta cell demise in both murine models and humans. The three aims of our R24 grant focus on defining how proinsulin folding patterns change when islets are challenged, and to identify how protein interactions with proinsulin may predict the efficiency of proinsulin trafficking through the secretory pathway, impacting islet health. Aim 1 will quantify the folded and unfolded state of proinsulin by measuring intermediates in the folding process in normal and diseased islets from well-characterized murine models. Aim 2 will define how the proteins that interact with proinsulin change during progression from normal, obese non-diabetic to T2D islets from human donors. Aim 3 will elucidate what interventions and chaperone functions may preserve productive proinsulin folding and restore an efficient proinsulin “proteostasis” network. Collectively, our proposed studies may identify novel biomarkers and avenues for therapeutic intervention in T2D, and therefore are of paramount importance to the mission of NIDDK.