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组基因之一，这些基因对β细胞功能至关重要，功能突变的丧失导致胰岛素不足导致永久性新生儿糖尿病。此外，Perk突变是男性先天性新生儿糖尿病最常见的原因。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完全不表达时，β细胞缺陷的严重程度和程度使得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。我们选择了另一种策略，即我们将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。该策略的一个关键假设是，更适度的表达变化可以改变生理结果。为了支持这一假设，我们已经证明，将Perk基因剂量减少一半会降低秋田胰岛素突变小鼠的血糖水平，并改善糖尿病的进展，而将Perk剂量增加到正常水平以上会导致血糖水平升高，并加速秋田小鼠糖尿病的进展。我们认为PERK是调节胰岛素原质量控制和内质网运输作为胰岛素生理需求的功能所必需的。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原胰岛素的质量控制和运输，确定PERK如何调节这些功能背后的ER伴随蛋白和折叠蛋白，并确定PERK活性的调节如何响应代谢的正常变化，从而介导胰岛素原运输和胰岛素分泌的变化，来验证这一假设。

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.