Physiological Regulation of Proinsulin Quality and Quantity Control in the Pancre

描述（由申请人提供）：本提案的重点是研究PERK eIF2a激酶（EIF2AK3）在调节内质网中胰岛素原运输和质量控制中的作用。PERK是在小鼠和人类中发现的一小部分基因之一，这些基因对β细胞功能至关重要，功能丧失突变会导致胰岛素不足导致永久性新生儿糖尿病。此外，Perk突变是人类近亲永久性新生儿糖尿病的最常见原因。我们对全球和组织特异性Perk KO小鼠的研究发现，PERK是正常β细胞增殖、胰岛素原运输和胰岛素分泌所必需的。然而，当PERK表达完全不存在时看到的β细胞缺陷的严重性和幅度使得Perk KO不适合作为发现β细胞中PERK的分子和生理功能的主要工具。我们选择了一种替代策略，我们将PERK表达从正常水平的0.2倍调节到2倍，然后研究对胰岛素原运输和胰岛素分泌的急性影响。这种策略的一个关键假设是，表达的更适度的变化可以改变生理结果。为了支持这一假设，我们已经表明，将Perk基因剂量减少一半导致血糖水平降低并改善秋田胰岛素突变小鼠的糖尿病进展，而将Perk剂量增加到高于正常水平导致血糖水平增加并加速秋田小鼠的糖尿病进展。我们建议，PERK是需要调节胰岛素原的质量控制和贩运的内质网作为一个功能的生理需求的胰岛素。因此，我们的总体策略是通过确定PERK如何调节野生型和秋田胰岛素原的质量控制和运输，确定PERK如何调节ER伴侣和折叠蛋白这些功能的基础，并确定PERK活性的调节如何响应于代谢的正常变化，以介导胰岛素原运输和胰岛素分泌的变化，来测试这一假设。

DESCRIPTION (provided by applicant): The focus of this proposal is to investigate the role of the PERK eIF2a kinase (EIF2AK3) in regulating proinsulin trafficking and quality control in the endoplasmic reticulum. PERK is among a small set of genes identified in mice and humans that are so critically important to beta cell functions that loss of function mutations result in permanent neonatal diabetes due to insulin insufficiency. Moreover, mutations in Perk are the most common cause of consanguineous permanent neonatal diabetes in man. Our investigation of global and tissue-specific Perk KO mice led to the discovery that PERK is required for normal beta cell proliferation, proinsulin trafficking, and insulin secretion. However, the severity and magnitude of the beta cell defects seen when PERK expression is entirely absent makes the Perk KO unsuitable as the primary tool to discover the molecular and physiological functions of PERK in beta cells. We have chosen an alternative strategy whereby we will modulate PERK expression from 0.2 to 2-fold of normal levels and then investigate the acute effects on proinsulin trafficking and insulin secretion. A key assumption of this strategy is that more modest changes in expression can alter physiological outcomes. In support of this assumption we have already shown that reducing Perk gene dosage in half results in reduced blood glucose levels and ameliorates the progression of diabetes of the Akita insulin mutant mouse, whereas increasing Perk dosage above normal results in increased blood glucose levels and accelerates the progression of diabetes in Akita mice. We propose that PERK is required to regulate proinsulin quality control and trafficking in the endoplasmic reticulum as a function of the physiological demand for insulin. Hence our overall strategy is to test the this hypothesis by determining how PERK regulates quality control and trafficking of wildtype and Akita proinsulin, determine how PERK regulates ER chaperone and folding proteins underlying these functions, and determine how the regulation of PERK activity responds to normal changes in metabolism to mediate changes in proinsulin trafficking and insulin secretion.