DeltaF508-CFTR Trafficking Regulated by 4-Phenylbutyrate

描述（由申请人提供）：囊性纤维化跨膜电导调节器 (CFTR) 是一种多功能蛋白质，既可将 Cl- 转运穿过上皮细胞的顶端质膜，又可调节其他蛋白质（例如上皮钠通道，ENaC）的离子转运。囊性纤维化 (CF) 的一个主要特征是气道上皮细胞中通过 ENaC 的 Na 转运过度活跃，尽管由于 CFTR 缺失而导致这种情况的机制尚不清楚。突变CFTR功能的药理学修复的努力，例如使用4-苯基丁酸钠(4PBA)来纠正最常见的突变CFTR、?F508-CFTR的运输，集中于评估突变CFTR的Cl-转运功能的恢复。这些研究常常忽略了 4PBA 或其他 CFTR“校正剂”对 ENaC 运输和/或功能的影响。我们提出的研究将解决药物修复或校正 ?F508-CFTR 运输是否会促进 CF 气道中 ENaC 的适当调节。这是实施改善?F508 CFTR 运输和功能的药理学策略的关键问题。我们的假设是，4PBA 诱导上皮细胞中分子伴侣表达的调节，从而导致 CFTR 和 ENaC 的细胞内运输和功能表达发生改变。由于 4PBA 还可以改善 CF 之外的疾病中多种突变蛋白的细胞内运输，因此这一假设也可能与开发针对多种其他蛋白质构象疾病的药物疗法密切相关。 4PBA 导致胞质伴侣 Hsc70 和 Hsp70 以及 29 kDa 的新型管腔内质网状蛋白 ERp29 的表达发生改变。我们的数据表明，这些伴侣蛋白的表达的特异性改变可调节 CFTR、?F508 和 ENaC 的运输和功能表达。本提案将建立在这些初步数据的基础上，并通过针对以下具体目标的研究来测试这一假设： 具体目标 1：确定 Hsc70 和 Hsp70 调节上皮细胞中 CFTR、F508 和 ENaC 运输的机制。具体目标 2：确定 ERp29（一种新型 4-苯基丁酸调节的管腔内质网状蛋白）调节上皮细胞中 CFTR、?F508 和 ENaC 运输的机制。 公众健康相关性：肺和气道保护自身免受环境影响的主要机制取决于呼吸道上皮细胞中适当的离子传输。这种离子传输在囊性纤维化中是异常的，导致显着的发病率和死亡率。这些数据将促进更好地了解负责囊性纤维化和其他气道疾病中离子运输的通道的调节，并为开发基于机制的囊性纤维化新疗法提供信息。

DESCRIPTION (provided by applicant): The Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) is a multifunctional protein that both transports Cl- across the apical plasma membrane of epithelial cells and regulates ion transport by other proteins, such as the Epithelial Sodium Channel, ENaC. A cardinal feature of Cystic Fibrosis (CF) is hyperactivity of Na+ transport via ENaC in the airway epithelia, although the mechanism by which this results from the absence of CFTR is not known. Efforts at pharmacologic repair of mutant CFTR function, such as the use of Sodium 4- Phenylbutyrate (4PBA) to correct trafficking of the most common mutant CFTR, ?F508-CFTR, have concentrated on assessing restoration of a mutant CFTR's Cl- transport function. These studies have often ignored the influence of 4PBA or other CFTR "correctors" on ENaC trafficking and/or function. Our proposed studies will address whether pharmacologic repair, or correction of ?F508-CFTR trafficking will promote appropriate regulation of ENaC in the CF airway. This is a key issue in the implementation of pharmacologic strategies to improve ?F508 CFTR trafficking and function. Our hypothesis is that 4PBA induces modulations of molecular chaperone expression in epithelial cells that result in altered intracellular trafficking and functional expression of CFTR and ENaC. Because 4PBA also improves the intracellular trafficking of a number of mutant proteins in diseases besides CF, this hypothesis may also be germane to developing pharmacologic therapies for a number of other protein conformational diseases. 4PBA causes altered expression of the cytosolic chaperones Hsc70 and Hsp70, as well as a novel luminal endoplasmic reticular protein of 29 kDa, ERp29. Our data suggest that specifically altered expression of these chaperones modulates CFTR, ?F508 and ENaC trafficking and functional expression. The present proposal will build on these preliminary data and test this hypothesis with studies directed at the following Specific Aims: Specific Aim 1: To determine the mechanism by which Hsc70 and Hsp70 modulate CFTR, ?F508 and ENaC trafficking in epithelial cells. Specific Aim 2: To determine the mechanism by which ERp29, a novel 4-phenylbutyrate-regulated luminal endoplasmic reticular protein, regulates the trafficking of CFTR, ?F508 and ENaC in epithelial cells. PUBLIC HEALTH RELEVANCE: The major mechanim by which the lung and airway defends itself from the environment depends on proper ion transport in the respiratory epithelia. Such ion transport is aberrant in Cystic Fibrosis, leading to significant morbidity and mortality. These data will promote better understanding of the regulation of channels responsible for ion transport in Cystic Fibrosis and other diseases of the airway, and inform development of novel, mechanism-based therapies for Cystic Fibrosis.