ERP29: PROMOTING ION CHANNEL BIOGENESIS FROM THE ER LUMEN

ERP29：促进 ER 腔的离子通道生物发生

• 批准号: 10302185
• 负责人: Ronald C Rubenstein
• 金额: $ 39.47万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10302185
• 关键词: ERP29; PROMOTING; ION; CHANNEL; BIOGENESIS

PROJECT SUMMARY/ABSTRACT Cystic Fibrosis (CF) and hypertension are life-shortening diseases of epithelial ion transport. In the CF airway, when the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) is functionally absent, there is a relative and unbalanced increase in the activity of the Epithelial Sodium Channel (ENaC) that leads to Na+ absorption from and depletion of the airway surface liquid. In turn, this impairs mucociliary clearance and predisposes the CF airway to bacterial colonization/infection. ENaC can also play a key role in causing hypertension; abnormally increased function of ENaC in the distal tubule of the nephron leads to increased Na+ retention, increased blood volume and hypertension. The regulation of ENaC activity is therefore key in understanding and developing therapies for these diseases of epithelial ion transport. A critical determinant of ENaC's open probability (Po), and therefore its function, is the proteolytic cleavage of the luminal/extracellular loops of its α and γ subunits; uncleaved channel has Po ~0, while fully cleaved channel has Po ~1. In fact, increased ENaC cleavage is reported in CF airway epithelial cells. Interestingly, ENaC can be delivered to the epithelial surface in either a cleaved or an uncleaved form, but the factor(s) that determine whether or not ENaC undergoes cleavage during biogenesis are not known. We have extensively studied Sodium 4-Phenylbutyrate (4PBA), the prototype small molecule corrector of the aberrant trafficking of the most common CF causing CFTR mutation, ΔF508. Others demonstrated that 4PBA also increases the function of ENaC. We found that ERp29 (Endoplasmic Reticulum Protein of 29 kD), a novel, ubiquitously expressed endoplasmic reticulum (ER) luminal chaperone has increased expression in bronchiolar epithelial cells treated with 4PBA, and that ERp29 promotes the trafficking of both wild type and ΔF508-CFTR. ERp29 was thus the first ER luminal component demonstrated to positively influence CFTR trafficking. We have also recently demonstrated that ERp29 promotes the function of ENaC. Interestingly, our data suggest that ERp29 does this by directing ENaC for cleavage in the Golgi during biogenesis, as depletion of ERp29 decreased ENaC function without altering expression of ENaC at the apical epithelial surface. The hypothesis of this proposal is that interaction of the luminal face of epithelial ion channels, such as CFTR and ENaC, with ERp29 during their biogenesis is required to direct these clients from the ER to the Golgi. This hypothesis will be tested with studies Specifically Aiming: 1) To test if ER-luminal facing mutations of CFTR that cause CF and are predicted to not interact with ERp29 have abnormal maturation. 2) To test the mechanism by which ERp29 directs ENaC to the Golgi during biogenesis. 3) To test the mechanism by which ENaC can bypass cleavage in the Golgi en route to the plasma membrane. These studies will yield key mechanistic insights that will be crucial in designing potential ERp29-directed therapeutic interventions for CFTR- and ENaC-opathies.

项目概要/摘要 囊性纤维化 (CF) 和高血压是 CF 中上皮离子转运的缩短寿命的疾病。 气道中，当囊性纤维化跨膜电导调节器（CFTR）功能缺失时， 上皮钠通道 (ENaC) 活性相对且不平衡的增加，导致 Na+ 气道表面液体的吸收和消耗反过来又会损害粘液纤毛的清除和吸收。 ENaC 也可能在引起 CF 气道细菌定植/感染方面发挥关键作用。 高血压；肾单位远端小管 ENaC 功能异常增加导致 Na+ 增加 因此，ENaC 活性的调节是关键。 了解并开发针对这些上皮离子转运疾病的疗法。 ENaC 开放概率 (Po) 及其功能的一个关键决定因素是 其 α 和 γ 亚基的管腔/细胞外环具有 Po ~0，而完全切割的通道 Po ~1 事实上，CF 气道上皮细胞中 ENaC 裂解增加。 以裂解或未裂解的形式递送至上皮表面，但决定因素 ENaC 在生物发生过程中是否经历裂解尚不清楚。 我们广泛研究了 4-苯基丁酸钠 (4PBA)，它是 最常见的 CF 的异常运输导致 CFTR 突变，ΔF508 其他人证明 4PBA。 我们发现 ERp29（29 kD 的内质网蛋白）也增强了 ENaC 的功能。 普遍表达的内质网 (ER) 管腔伴侣在细支气管中表达增加 用 4PBA 处理的上皮细胞，ERp29 促进野生型和 ΔF508-CFTR 的运输。 因此，ERp29 是第一个被证明可以对 CFTR 运输产生积极影响的 ER 管腔成分。 我们最近还证明 ERp29 可以促进 ENaC 的功能。 表明 ERp29 通过指导 ENaC 在生物发生过程中在高尔基体中裂解来实现这一点，因为 ERp29 降低 ENaC 功能，但不改变根尖上皮表面 ENaC 的表达。 该提议的假设是上皮离子通道的管腔面的相互作用，例如 CFTR 和 ENaC 在其生物发生过程中与 ERp29 一起需要将这些客户从 ER 引导至高尔基体。 假设将通过研究进行检验 具体目的： 1) 测试 CFTR 的 ER 腔面突变是否会导致 导致 CF 且预计不与 ERp29 相互作用的成熟异常 2) 通过以下方式测试该机制。 ERp29 在生物发生过程中将 ENaC 引导至高尔基体 3) 测试 ENaC 绕过的机制。 这些研究将产生关键的机制见解。 对于设计针对 CFTR 和 ENaC 疾病的潜在 ERp29 导向治疗干预措施至关重要。

项目成果

Bitter taste receptor agonists regulate epithelial two-pore potassium channels via cAMP signaling.

• DOI: 10.1186/s12931-021-01631-0
• 发表时间: 2021-01-28
• 期刊: Respiratory research
• 影响因子: 5.8
• 作者: Kohanski MA;Brown L;Orr M;Tan LH;Adappa ND;Palmer JN;Rubenstein RC;Cohen NA
• 通讯作者: Cohen NA

Editorial Focus: CFTR-dependent bicarbonate secretion by Calu-3 cells.

• DOI: 10.14814/phy2.13691
• 发表时间: 2018-05
• 期刊: Physiological reports
• 影响因子: 2.5
• 作者: Rubenstein RC

ERp29 as a regulator of Insulin biosynthesis.

ERp29 作为胰岛素生物合成的调节剂。

• DOI: 10.1371/journal.pone.0233502
• 发表时间: 2020
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Viviano,Jeffrey;Brecker,Margaret;Ferrara-Cook,Christine;Suaud,Laurence;Rubenstein,RonaldC
• 通讯作者: Rubenstein,RonaldC

Correction: ERp29 as a regulator of Insulin biosynthesis.

更正：ERp29 作为胰岛素生物合成的调节剂。

• DOI: 10.1371/journal.pone.0238264
• 发表时间: 2020
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Viviano,Jeffrey;Brecker,Margaret;Ferrara-Cook,Christine;Suaud,Laurence;Rubenstein,RonaldC
• 通讯作者: Rubenstein,RonaldC

Safety and efficacy of treatment with lumacaftor in combination with ivacaftor in younger patients with cystic fibrosis.

lumacaftor 与 ivacaftor 联合治疗年轻囊性纤维化患者的安全性和有效性。

• DOI: 10.1080/17476348.2019.1602040
• 发表时间: 2019
• 期刊: Expert review of respiratory medicine
• 影响因子: 3.9
• 作者: Cheng,PiChun;Alexiou,Stamatia;Rubenstein,RonaldC
• 通讯作者: Rubenstein,RonaldC