DeltaF508-CFTR Trafficking Regulated by 4-Phenylbutyrate

描述（由申请人提供）：囊性纤维化跨膜传导调节剂（CFTR）是一种多功能蛋白，既可以通过上皮细胞的顶质膜运输Cl-，也可以通过其他蛋白（如上皮钠通道，ENaC）调节离子运输。囊性纤维化（CF）的一个主要特征是气道上皮中通过ENaC的Na+转运过度活跃，尽管CFTR缺失导致这一现象的机制尚不清楚。在药物修复突变CFTR功能方面的努力，如使用4-苯基丁酸钠（4PBA）来纠正最常见的CFTR突变体的贩运，？F508-CFTR，主要研究CFTR突变体Cl-转运功能的恢复。这些研究往往忽略了4PBA或其他CFTR“校正剂”对ENaC转运和/或功能的影响。我们提出的研究将探讨药物修复或纠正？F508-CFTR的转运将促进CF气道内ENaC的适当调节。这是一个关键问题，在实施药理学策略，以提高？F508 CFTR的流量和功能。我们的假设是，4PBA诱导了上皮细胞中分子伴侣表达的调节，从而改变了细胞内运输和CFTR和ENaC的功能表达。因为除了CF外，4PBA还可以改善许多突变蛋白在疾病中的细胞内运输，所以这一假设也可能与开发许多其他蛋白质构象疾病的药物治疗密切相关。4PBA导致细胞质伴侣Hsc70和Hsp70以及一种29 kDa的新型内质网状蛋白ERp29的表达改变。我们的数据表明，这些伴侣蛋白的特异性表达改变可调节CFTR。F508与ENaC的转染及功能表达。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验证这一假设：具体目标1：确定Hsc70和Hsp70调节CFTR的机制；上皮细胞中F508和ENaC的转运。特异性目标2：确定ERp29（一种新型4-苯基丁酸调节的腔内质网状蛋白）调节CFTR运输的机制。上皮细胞中的F508和ENaC。

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.