COBRE: UNR: MOLECULAR IDENTIF & CHAR OF CALCIUM-ACTIVATED CHLORIDE CHANNELS

COBRE：UNR：分子鉴定

• 批准号: 7381163
• 负责人: FIONA Catherine BRITTON
• 金额: $ 22.72万
• 依托单位: UNIVERSITY OF NEVADA RENO
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-06-01 至 2007-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7381163
• 关键词: COBRE; UNR; MOLECULAR; IDENTIF; CHAR

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Molecular biological studies of ion channels have revolutionized our perception of the structure and mechanism of action of these physiologically critical proteins. Any comprehensive examination of anion channels in the cardiovascular system must include a goal of identification and characterization of the molecular components underlying these conductances.We have made great strides in recent years towards the goal of molecular identification of cardiac Cl- channels. We have identified ICl.cAMP, ICl.PKC and ICl.ATP as being encoded by a splice variant of CFTR; we have identified ICl.vol as ClC-3; we have preliminary data for a relationship between ICl.ir and ClC-2. However, the gene(s) that code for the ion channel underlying a key cardiovascular chloride current (ICl.Ca) remains to be determined. This Center of Biomedical Research Excellence will provide the tools and the collaborative resources required to address this question. We will test our general hypothesis that ICl.Ca in cardiac myocytes is encoded by a member(s) of the CLCA and/or novel bestrophin gene family. We propose to (1) Determine the expression pattern of molecular forms of bestrophins and how that data relates to the distribution of native ICl.Ca. RT-PCR and nuclease protection studies will determine if novel splice variants are expressed in cardiovascular tissues. (2) Determine the biophysical properties of cloned cardiac CLCA and bestrophin genes functionally expressed in mammalian cells lines and evaluate their relationship to native I.ClCa. (3) Determine the immunolocalization of bestrophins at tissue and cellular levels to determine cardiovascular cell types expressing bestrophin channels and to investigate their co-localization with each other. (4) Determine the physiological role bestrophin proteins play in I.ClCa in heart by eliminating bestrophin protein expression in an inducible heart-specific knockout mouse.

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。离子通道的分子生物学研究彻底改变了我们对这些生理上至关重要的蛋白质的结构和作用机制的认识。任何对心血管系统阴离子通道的全面检查都必须包括鉴定和表征这些传导的分子成分的目标。近年来，我们在心脏Cl-通道分子鉴定方面取得了很大的进展。我们已经确认了ICl。营地,ICl。PKC和ICl。ATP由CFTR的剪接变体编码；我们已经确认了ICl。vol为ClC-3；我们有ICl之间关系的初步数据。ir和ClC-2。然而，编码关键心血管氯离子电流（ICl.Ca）的离子通道的基因仍有待确定。这个卓越生物医学研究中心将提供解决这个问题所需的工具和协作资源。我们将检验我们的一般假设ICl。心肌细胞中的Ca是由CLCA和/或新的肌营养蛋白基因家族的成员编码的。我们建议：(1)确定strophins分子形式的表达模式以及该数据与天然ICl.Ca分布的关系。RT-PCR和核酸酶保护研究将确定新的剪接变异是否在心血管组织中表达。(2)确定克隆的心脏CLCA和bestrophin基因在哺乳动物细胞系中功能表达的生物物理特性，并评估其与天然I.ClCa的关系。(3)测定besstrophin在组织和细胞水平的免疫定位，以确定表达besstrophin通道的心血管细胞类型，并研究它们之间的共定位。(4)通过在可诱导的心脏特异性敲除小鼠中消除bestrophin蛋白的表达，确定bestrophin蛋白在心脏i - clca中的生理作用。