IDENTIFICATION OF CA2+ REGULATED APICAL CL- CHANNELS

CA2 调节的顶端 CL 通道的识别

基本信息

批准号：
6638538
负责人：
SHERIF E GABRIEL
金额：
$ 21.68万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-04-01 至 2005-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6638538
关键词：
apical membrane biological signal transduction calcium flux cell line chloride channels cystic fibrosis gene mutation molecular cloning protein isoforms protein structure function respiratory epithelium voltage /patch clamp

项目摘要

DESCRIPTION (Adapted from applicant's abstract): Cystic fibrosis (CF) is characterized as a defect of electrolyte transport of epithelial cells of exocrine tissues. Central to the pathophysiology of CF is the absence of cAMP- stimulated Cl- secretion and an enhanced rate of Na+ absorption. The cloning of the CF gene and characterization of the gene product, the cystic fibrosis transmembrane conductance regulator (CFTR), has revealed that CFTR functions as a cAMP-stimulated Cl- channel. Cloning of the CF gene led to the creation of an animal model for CF, the CFTR (-/-) knockout mouse. Interestingly, the CFTR (-/-) mouse did not exhibit a severe CF phenotype in many epithelial tissues, including the airways. Importantly, the absence of a CF phenotype led to the formal identification of the Ca2+-regulated, or "alternative" Cl- conduction pathway (Cl-A) that is molecularly distinct from CFTR and plays a protective role in preventing CF pathogenesis in the airways of the CFTR (-/-) mouse. This proposal focuses on Cl-A and hypothesizes that Cl-A is an important airway epithelial ion channel resident in the apical membrane and regulated by elevation of intracellular Ca2+. Cl-A will be investigated at three levels, 1) characterization of transepithelial Cl- currents, 2) regulation of Cl-A by Ca2+-mediated signal transduction pathways, 3) patch clamp identification of the Cl-A single channel properties. Confluent polarized epithelial preparations will be used to characterize the Ca2+-stimulated Cl- current and to identify the agonists and signal transducers that regulate this conductance. Patch Cl-Amp technique will be used to identify the Cl-A single channel properties that correlate with Ca2+-stimulated currents from CF murine airway epithelial cells. Importantly all single channel studies will be performed on cells grown on a permeablized support and only channels resident in the apical membrane will be studied. Finally, comparisons between the endogenous Cl-A and heterologously expressed candidate clones will permit the unequivocal identification of the Ca2+-activated airway Cl- channel. A complete characterization of Cl-A will lead to the genesis of new therapies for CF disease that would circumvent a defective CFTR.

描述（改编自申请人的摘要）：囊性纤维化（CF）是被描述为上皮细胞电解质转运的缺陷外分泌组织。 CF病理生理学的中心是没有营地刺激的Cl-分泌和Na+吸收的速率增强。克隆 CF基因和基因产物的表征，囊性纤维化跨膜电导调节剂（CFTR）表明，CFTR的功能为一个营地刺激的Cl-通道。 CF基因的克隆导致创建 CF的动物模型，CFTR（ - / - ）基因敲除鼠标。有趣的是，CFTR （ - / - ）小鼠在许多上皮组织中没有表现出严重的CF表型，包括航空公司。重要的是，缺乏CF表型导致 CA2+调节或“替代” Cl-传导的形式识别途径（Cl-A）与CFTR分子不同并具有保护性在CFTR（ - / - ）小鼠气道中预防CF发病机理中的作用。这提案重点是CL-A，并假设CL-A是重要的气道上皮离子通道驻留在顶膜中，并受到调节细胞内Ca2+的升高。 CL-A将在三个级别进行研究，1） transpithelial cl currents的表征，2）CL-A调节 Ca2+介导的信号转导途径，3）贴片夹的识别 CL-A单个通道属性。汇合极化上皮制剂将用于表征Ca2+刺激的Cl-电流并识别调节该电导的激动剂和信号传感器。修补 CL-AMP技术将用于识别CL-A单个通道属性与CF鼠气上皮的Ca2+刺激电流相关细胞。重要的是，所有单个通道研究都将在生长的细胞上进行在透化的支撑和仅驻留在顶膜中的通道将被研究。最后，内源性CL-A和异源表达的候选克隆将允许明确 CA2+活化的气道CL-通道的识别。完整 CL-A的表征将导致CF新疗法的起源疾病会避免有缺陷的CFTR。

项目成果

期刊论文数量（4）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Regulation of murine airway surface liquid volume by CFTR and Ca2+-activated Cl- conductances.

DOI：
10.1085/jgp.20028599
发表时间：
2002-09
期刊：
The Journal of general physiology
影响因子：
0
作者：
Tarran R;Loewen ME;Paradiso AM;Olsen JC;Gray MA;Argent BE;Boucher RC;Gabriel SE
通讯作者：
Gabriel SE

Apical localization of ITPK1 enhances its ability to be a modifier gene product in a murine tracheal cell model of cystic fibrosis.

ITPK1 的顶端定位增强了其在囊性纤维化小鼠气管细胞模型中作为修饰基因产物的能力。