Perk eIF2a Kinase Integrates Proinsulin Quality Control and Insulin Secretion

 描述（由申请方提供）：永久性新生儿糖尿病和2型β细胞失代偿的单基因形式突出了内质网（ER）中胰岛素原加工的重要性和脆弱性。胰岛素原的折叠、ER相关蛋白降解（ERAD）介导的质量和数量控制以及胰岛素原从ER向高尔基体的运输等过程一度被认为是管家功能，但这些过程都受到代谢状态和胰岛素需求的高度调节。我们已经确定PERK eIF 2a激酶，其功能丧失导致沃尔科特Rallison综合征中的永久性新生儿糖尿病，作为ER中胰岛素原加工的关键调节因子。我们假设了一个新的和有争议的模型，PERK调节胰岛素原的加工通过控制关键ER分子伴侣和折叠蛋白，包括GRP 78和ERp 72，这反过来又控制胰岛素原是否被ERAD途径降解或贩运到高尔基体进行蛋白水解加工和包装成分泌颗粒的表达。在本提案中，我们将通过操纵PERK-eIF 2a通路和关键ER分子伴侣的表达并评估胰岛素原的命运和加工状态来进一步测试该模型。我们还将确定PERK-eIF 2a如何调节胰岛素原加工以响应波动的葡萄糖水平，从而提供适当水平的胰岛素产生。基于Guanabenz，一种eIF 2a去磷酸化抑制剂，可以逆转在PERK缺陷型β细胞中观察到的严重胰岛素原聚集的初步结果，我们提出测试其预防或逆转与沃尔科特Rallison综合征和显性负作用胰岛素突变体（MIDY）相关的严重永久性新生儿糖尿病的能力。该策略也可适用于治疗与2型糖尿病失代偿相关的胰岛β细胞功能障碍。最终目的是确定ER分子伴侣的PERK依赖性调节的分子机制，并且对于理解PERK在调节ER细胞功能中发挥核心作用的调节背景是必不可少的。

 DESCRIPTION (provided by applicant): Monogenic forms of permanent neonatal diabetes and type 2 ß-cell decompensation have highlighted the importance and vulnerability of proinsulin processing in the endoplasmic reticulum (ER). Once thought to be housekeeping functions, the processes of proinsulin folding, ER associated protein degradation (ERAD)- mediated quality and quantity controand trafficking of proinsulin from the ER to the Golgi are all highly regulated as a function of the metabolic state and demand for insulin. We have identified the PERK eIF2a kinase, whose loss of function results in permanent neonatal diabetes in the Wolcott Rallison syndrome, as a key regulator of proinsulin processing in the ER. We postulated a novel and controversial model that PERK regulates proinsulin processing by controlling the expression of key ER chaperone and folding proteins including GRP78 and ERp72, which, in turn, control whether proinsulin is degraded by the ERAD pathway or trafficked forward to the Golgi for proteolytic processing and packaging into secretory granules. In this proposal we will further test this model by manipulating the PERK-eIF2a pathway and the expression of key ER chaperones and assessing the fate and processing state of proinsulin. We will also determine how PERK- eIF2a modulates proinsulin processing in response to fluctuating glucose levels in order to provide the appropriate level of insulin production. Based on preliminary results that Guanabenz, an eIF2a dephosphorylation inhibitor, can reverse the severe proinsulin aggregation seen in PERK deficient ß-cells, we proposed to test its ability to prevent or reverse the severe permanent neonatal diabetes associated with the Wolcott Rallison Syndrome and dominant-negative acting insulin mutants (MIDY). This strategy may also be applicable to treatment of ß-cell dysfunctions associated with type 2 diabetes decompensation. The final Aim is designed to identify the molecular mechanisms underlying PERK-dependent regulation of ER chaperones and is essential for understanding the regulatory context in which PERK plays a central role in regulating ß-cell functions.