Functional Relevance of CFTR Trafficking In Vivo to Intestinal Disease

CFTR 体内贩运与肠道疾病的功能相关性

基本信息

批准号：
9079459
负责人：
Nadia A. Ameen
金额：
$ 36.21万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-04-01 至 2018-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9079459
关键词：
Acidity Anions Apical Bicarbonates Binding Cells Chloride Channels Chloride Ion Chlorides Chronic Constipation Cyclic Nucleotides Cystic Fibrosis Cystic Fibrosis Transmembrane Conductance Regulator Cytoplasm Diarrhea Disease Drug usage Endocytosis Endosomes Enterocytes Enterotoxins Epithelial Epithelium Equilibrium Exocytosis Failure Figs - dietary Fluids and Secretions Functional disorder Grant Health Intestinal Diseases Intestinal Obstruction Intestines Investigation Knockout Mice Knowledge Laboratories Lead Link Liquid substance Membrane Molecular Molecular Motors Motor Movement Myosin ATPase Pathway interactions Physiological Population Proteins Publishing Recycling Regulation Reporting Retrieval Role Second Messenger Systems Site Small Intestines Surface TFAP2A gene Testing Tissues Transcription Factor AP-2 Alpha Villus acute stress apical membrane biological adaptation to stress brush border membrane cell type density fluidity in vivo insight mouse model mutant myosin VI response second messenger therapy development trafficking

项目摘要

DESCRIPTION (provided by applicant): Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) channels are the major apical exit pathways for anion (chloride and bicarbonate) secretion from the brush border membrane (BBM) in the intestine. Absence of CFTR on the enterocyte BBM leads to hyperacidity and intestinal obstruction as observed in the disease Cystic Fibrosis (CF). Increased CFTR on the BBM leads to secretory diarrhea. Our studies identified a unique subcellular distribution for CFTR in the intestine that is important fo both basal and increased fluid secretion under pathophysiologic conditions such as bacterial enterotoxins and low luminal pH. Unlike other epithelia that express CFTR only on the BBM, enterocytes express CFTR both on the BBM and subapical endosomes. The BBM fraction maintains basal intestinal fluidity by constitutive recycling of CFTR. The larger fraction of CFTR in subapical endosomes is important for rapid stress responses. For example, bacterial enterotoxins increase intracellular cyclic nucleotides that lead to rapid insertion of endosomal CFTR into the BBM to increase the number of CFTR channels and elicit massive fluid secretion. Similarly, luminal acidity elicits robust CFTR traffic into the BBM of villus enterocytes in the smll intestine to secrete bicarbonate. Thus, luminal pH in the CF small intestine is acidic. The protein motors and factors that direct CFTR traffic into and out of the enterocyte BBM are not known and what is known is largely credited to this laboratory. Previous investigations revealed that a minus motor Myosin6 (Myo6) directs CFTR endocytosis from the BBM of crypt and villus enterocytes. During the previous R01 cycle, the specific adaptor (AP-2-�) that links CFTR to the Myo6 motor was identified. Proposed studies will investigate the site of CFTR- AP-2� binding, elucidate the mechanism and site of action of AP-2� in the intestine. Myo6 operates in tandem with its counterpart plus end motor Myosin 1a (Myo1a) in the enterocyte. Proposed studies in the first application sought to investigate the role of Myo1a in CFTR BBM localization and function in Myo1a KO mice. These studies were published. Myo1a is the exocytic motor for CFTR traffic from endosomes into the BBM of villus enterocytes. The mechanisms regulating Myo1a-dependent exocytosis of CFTR will now be examined. New studies in Aim 3 will use a Myo6/Myo1a double KO mouse model to test the hypothesis that Myo6 and Myo1a function as counterpart motors to maintain proper CFTR BBM localization and function by endocytic and exocytic traffic into and out of the enterocyte BBM. Proposed studies will expand critical knowledge gained during the previous cycle on factors and mechanisms regulating CFTR in the intestine under homeostatic and pathophysiologic conditions and extend insights into CFTR trafficking mechanisms that can be targeted to treat common intestinal diseases such as constipation.

描述（由申请人提供）：囊性纤维化跨膜电导调节器 (CFTR) 通道是肠内刷状缘膜 (BBM) 分泌阴离子（氯离子和碳酸氢根）的主要顶端出口途径。肠上皮细胞 BBM 引线上不存在 CFTR。囊性纤维化 (CF) 疾病中观察到的胃酸过多和肠梗阻 BBM 上的 CFTR 增加会导致分泌。我们的研究发现了 CFTR 在肠道中的独特亚细胞分布，这对于细菌肠毒素和低腔 pH 等病理生理条件下的基础液体分泌和增加液体分泌都很重要，与其他仅在 BBM 上表达 CFTR 的上皮细胞不同，肠细胞表达 CFTR。 BBM 和根尖下内体上的 BBM 部分通过 CFTR 的组成型再循环维持肠道流动性。内体对于快速应激反应很重要，例如，细菌肠毒素会增加细胞内的环核苷酸，从而导致内体 CFTR 快速插入 BBM，从而增加 CFTR 通道的数量并引发大量液体分泌。小肠绒毛肠细胞的 BBM 分泌碳酸氢盐，因此，CF 小肠内的管腔 pH 值呈酸性。引导 CFTR 进出肠细胞 BBM 的马达和因素尚不清楚，已知的大部分归功于该实验室之前的研究表明，负运动肌球蛋白 6 (Myo6) 指导隐窝和绒毛肠细胞 BBM 的 CFTR 内吞作用。在之前的 R01 周期中，确定了将 CFTR 连接到 Myo6 电机的特定适配器 (AP-2-�)。拟议的研究将调查 CFTR- 的位点。 AP-2� 结合，阐明了 AP-2� 在肠道中与其对应物加末端运动肌球蛋白 1a (Myo1a) 协同作用的机制和作用位点。第一个应用中提出的研究旨在调查。 Myo1a 在 Myo1a KO 小鼠 CFTR BBM 定位和功能中的作用这些研究已发表，Myo1a 是 CFTR 从内体运输到体内的胞吐马达。现在将检查调节 CFTR 的 Myo1a 依赖性胞吐作用的机制，Aim 3 中的新研究将使用 Myo6/Myo1a 双 KO 小鼠模型来测试 Myo6 和 Myo1a 作为对应马达以维持正常状态的假设。 CFTR BBM 通过进出肠细胞 BBM 的内吞和胞吐运输进行定位和功能。拟议的研究将扩展上一周期中获得的有关因素和因素的关键知识。体内平衡和病理生理条件下调节肠道 CFTR 的机制，并深入了解可用于治疗便秘等常见肠道疾病的 CFTR 运输机制。