Gap Junction Trafficking to and within the Plasma Membrane

间隙连接运输到质膜和质膜内

• 批准号: 7806535
• 负责人: Robin M Shaw
• 金额: $ 38.63万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-15 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7806535
• 关键词: Actins; Affect; Arrhythmia; Behavior; Binding; Biochemistry; Blood flow; Cardiac; Cardiac Myocytes; Cell membrane; Cells; Cellular biology; Clinical; Congestive Heart Failure; Connexin 43; Connexon; Coupling; Cytoskeleton; Data; Diffuse; Diffusion; Disease; Elements; Event; Fluorescence Microscopy; Functional disorder; Future; Gap Junctions; Goals; Growth; Heart failure; Imaging Techniques; Imaging technology; Intercalated disc; Ion Channel; Ischemia; Knowledge; Laboratories; Lateral; Life; Location; Measures; Membrane; Microtubules; Molecular; Morbidity - disease rate; Movement; Myocardial Ischemia; Oxidative Stress; Pattern; Positioning Attribute; Process; Property; Proteins; Regulation; Resolution; Role; Simulate; Stress; Sudden Death; Techniques; Testing; Time; United States; Ventricular; Work; base; cellular imaging; charge coupled device camera; fluorescence imaging; heart cell; immunocytochemistry; inhibitor/antagonist; mortality; prevent; protein transport; public health relevance; sudden cardiac death; targeted delivery; therapeutic target; therapy development; trafficking; Actins; Affect; Arrhythmia; Behavior; Binding; Biochemistry; Blood flow; Cardiac; Cardiac Myocytes; Cell membrane; Cells; Cellular biology; Clinical; Congestive Heart Failure; Connexin 43; Connexon; Coupling; Cytoskeleton; Data; Diffuse; Diffusion; Disease; Elements; Event; Fluorescence Microscopy; Functional disorder; Future; Gap Junctions; Goals; Growth; Heart failure; Imaging Techniques; Imaging technology; Intercalated disc; Ion Channel; Ischemia; Knowledge; Laboratories; Lateral; Life; Location; Measures; Membrane; Microtubules; Molecular; Morbidity - disease rate; Movement; Myocardial Ischemia; Oxidative Stress; Pattern; Positioning Attribute; Process; Property; Proteins; Regulation; Resolution; Role; Simulate; Stress; Sudden Death; Techniques; Testing; Time; United States; Ventricular; Work; base; cellular imaging; charge coupled device camera; fluorescence imaging; heart cell; immunocytochemistry; inhibitor/antagonist; mortality; prevent; protein transport; public health relevance; sudden cardiac death; targeted delivery; therapeutic target; therapy development; trafficking

DESCRIPTION (provided by applicant): Ischemic heart disease and its associated complications of sudden cardiac death and congestive heart failure remain leading causes of morbidity and mortality in the United States. The arrhythmogenic pathophysiology of ischemic disease includes loss of cardiomyocyte electrical coupling through altered localization and modulation of gap junctions. The precise molecular mechanisms underlying altered coupling remain elusive. The objective of this proposal is to understand the cellular movement of gap junction hemichannels (connexons) in normal and ischemic conditions. Our central hypothesis is that connexons require the cytoskeleton to target them to specific locations on the plasma membrane and, once in the plasma membrane, there is a limited role of lateral diffusion and other means of non- cytoskeleton based channel movement. The hypothesis will be tested by using fixed and live cell imaging techniques including high resolution total internal reflection fluorescence (TIRF) imaging, with supplementary biochemistry, to understand the molecular mechanisms of connexon trafficking. Particular aims include understanding the role of microtubule based directed targeting of connexons to intercalated discs in conditions of oxidative stress and simulated ischemia; to understand the role of the actin cytoskeleton in targeted delivery of connexons; and to determine quantitatively the capacity of connexons to diffuse laterally to other membrane regions within the plasma membrane. Preliminary data indicate that oxidative stress limits microtubule capture of cortical membrane, preventing delivery of connexons to the plasma membrane; that actin helps microtubules position and target ion channels to specific regions of the cortical membrane, and that lateral diffusion of connexons is highly restricted. These aims will further our understanding of the regulation and behavior of cardiac gap junctions. The general field of protein trafficking will benefit from fundamental new knowledge about cytoskeleton and ischemic type regulation of these channels. Furthermore, key molecules and events in the trafficking of gap junctions will be identified to be used as therapeutic targets to lessen the arrhythmias and dysfunction associated with ischemic heart disease. PUBLIC HEALTH RELEVANCE: The clinical sequelae of ischemic heart disease are congestive heart failure and sudden cardiac death which are primary causes of mortality in the United States. The cellular basis of both heart failure and sudden death involve diminished electrical coupling between heart cells. This application proposes to study the molecular mechanisms of electrical coupling between heart cells and identify proteins involved in regulating the coupling under normal conditions and during times of reduced blood flow (ischemia).

描述（由申请人提供）：缺血性心脏病及其相关心脏死亡和充血性心力衰竭的并发症仍然是美国发病和死亡率的主要原因。缺血性疾病的心律失常性病理生理学包括通过改变缝隙连接的定位和调节来丧失心肌细胞电耦合。耦合改变的确切分子机制仍然难以捉摸。该建议的目的是了解正常和缺血性条件下间隙连接半通道（连接）（连接）的细胞运动。我们的中心假设是，连接子要求细胞骨架将它们靶向质膜上的特定位置，并且在质膜中，侧向扩散和其他非细胞骨架基基通道运动的作用有限。该假设将通过使用固定和活细胞成像技术进行检验，包括高分辨率的总内部反射荧光（TIRF）成像，并具有补充生物化学，以了解康纳克斯子运输的分子机制。特定目的包括了解在氧化应激和模拟缺血条件下，基于微管的指示靶向连接盘的作用；了解肌动蛋白细胞骨架在靶向输送连接子中的作用；并定量确定连接子横向扩散到质膜内其他膜区域的能力。初步数据表明，氧化应激限制了皮质膜的微管捕获，从而阻止了连接膜向质膜递送。该肌动蛋白有助于微管位置和靶向离子通道到皮质膜的特定区域，并且高度限制了连接子的侧向扩散。这些目标将进一步理解心脏间隙连接的调节和行为。蛋白质运输的一般领域将受益于有关这些通道的细胞骨架和缺血性调节的基本知识。此外，将确定缝隙连接的运输中的关键分子和事件被确定为减少与缺血性心脏病相关的心律不齐和功能障碍的治疗靶标。公共卫生相关性：缺血性心脏病的临床后遗症是充血性心力衰竭和心脏猝死，这是美国死亡率的主要原因。心力衰竭和猝死的细胞基础涉及心脏细胞之间的电耦合减少。该应用建议研究心脏细胞之间电耦合的分子机制，并鉴定在正常条件下和血液流量降低（缺血）下涉及调节偶联的蛋白质。