Molecular Determinants of Function in Kir2.x channels

Kir2.x 通道功能的分子决定因素

• 批准号: 7924588
• 负责人: JUSTUS M ANUMONWO
• 金额: $ 29.06万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2012-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7924588
• 关键词: Arrhythmia; Binding; Biochemical; Biological; Biophysics; C-terminal; Cardiac; Cardiac Myocytes; Cell membrane; Cell surface; Cells; Consensus; Consensus Sequence; Cyclic AMP-Dependent Protein Kinases; DLG1 gene; Genes; Heart; Human; In Vitro; Incidence; Intercalated disc; Ion Channel; Lead; Macromolecular Complexes; Membrane; Modification; Molecular; Myocardium; Neonatal; Paralysed; Phosphorylation; Play; Probability; Property; Proteins; Recruitment Activity; Regulation; Research Personnel; Role; Scaffolding Protein; Sorting - Cell Movement; Surface; Surface Plasmon Resonance; Syndrome; Techniques; Tertiary Protein Structure; Testing; base; in vivo; loss of function; programs; protein protein interaction; research study; synapse-associated protein 97; three dimensional structure; trafficking

DESCRIPTION (provided by applicant): The inward rectifier current (IK1) plays a role in the excitability properties of the cardiac cell. Loss of function in one of the genes (KCNJ2) underlying the channel protein (Kir2.1) in humans leads to Andersen's syndrome, characterized by dysmorphic features, paralysis and cardiac arrhythmias. Properties of ion channels depend on their three dimensional structure as well as by their interactions with accessory proteins. There is little information available on proteins that directly associate with Kir2.x channels, or how these proteins modify channel function in the heart. The scaffolding proteins, (SAP97, Veli and CASK) that target, cluster and regulate ion channels, have been shown to directly bind or are co localized with Kir2.x channels in the heart, with implications that are largely unknown. Our objective is to determine the protein-protein interactions involved in the regulation of Kir2.x channel function. Our main hypothesis is that SAP97, Veli and CASK interactions with Kir2.x channels results in modification of channel biophysics, regulation, expression and subcellular targeting in the heart. We will use a combination of approaches including electrophysiological, surface plasmon resonance, biochemical and molecular biological techniques and carry out our studies in HEK293 cells and in isolated cardiac myocytes. Specific Aims: 1) To investigate the molecular basis of changes in the biophysical properties of Kir2.x channels following co-expression with cardiac PDZ domain proteins. 2) To study the impact of interactions between Kir2.x channels and PDZ domain proteins on channel phosphorylation by PKA. 3) To determine the role of PDZ domain proteins on Kir2.x channel expression, localization and function in cardiac and non-cardiac cells. These studies should lead to a better understanding of the molecular properties that modulate the function of the classical cardiac inward rectifier current.

描述（由申请人提供）：内向整流器电流（IK1）在心脏细胞的兴奋性特性中起作用。人类通道蛋白（KIR2.1）的基因之一（KCNJ2）中功能的丧失导致安德森综合征，其特征是畸形特征，瘫痪和心律不齐。离子通道的性质取决于其三维结构以及与附件蛋白的相互作用。直接与KIR2.X通道直接关联的蛋白质的信息很少，或者这些蛋白质如何修饰心脏中的通道功能。靶向，簇和调节离子通道的脚手架蛋白（SAP97，Veli和Cask）已被证明直接结合或与心脏中的Kir2.x通道一起定位，其含义在很大程度上是未知的。我们的目标是确定与KIR2.x通道功能调节有关的蛋白质蛋白质相互作用。我们的主要假设是，SAP97，Veli和Cask与Kir2.x通道的相互作用会导致心脏中通道生物物理学，调节，表达和亚细胞靶向的改变。我们将结合多种方法，包括电生理，表面等离子体共振，生化和分子生物学技术，并在HEK293细胞和孤立的心肌细胞中进行研究。具体目的：1）研究与心脏PDZ结构蛋白共表达后，Kir2.x通道生物物理特性变化的分子基础。 2）研究KIR2.x通道和PDZ结构蛋白之间相互作用对PKA通道磷酸化的影响。 3）确定PDZ结构蛋白对心脏和非心脏细胞中KIR2.x通道表达，定位和功能的作用。这些研究应更好地理解调节经典心脏内向整流器电流功能的分子特性。