Molecular Structure and Regulation of the Permeability Transition Pore

渗透率转变孔的分子结构和调控

• 批准号: 8370446
• 负责人: MICHAEL A FORTE
• 金额: $ 32.39万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8370446
• 关键词: Address; Alzheimer' s Disease; Amyotrophic Lateral Sclerosis; Benzodiazepine Receptor; Biochemical Genetics; Brain; Cell Death; Collagen; Databases; Disease; Elements; Event; Funding; Future; Generations; Genetic; Genetic screening method; Goals; Heart; Homeostasis; Human; Hydrogen Peroxide; In Vitro; Inner mitochondrial membrane; Knock-out; Knowledge; Laboratories; Lead; Mitochondria; Mitochondrial Proteins; Modeling; Molecular; Molecular Structure; Morphologic artifacts; Multiple Sclerosis; Muscular Dystrophies; Myocardial Infarction; Outcome; Oxidative Stress; Oxidoreductase; Pathology; Pathway interactions; Perception; Peripheral; Permeability; Physiological; Play; Pore Proteins; Process; Property; Protein Isoforms; Proteins; Proteome; Proteomics; Protocols documentation; Regulation; Reperfusion Injury; Repression; Resistance; Role; Staging; Stroke; Structure; Testing; Work; base; design; expectation; human disease; in vivo; interest; mitochondrial dysfunction; mitochondrial permeability transition pore; mouse model; novel; novel strategies; novel therapeutics; porin; protein expression; response; therapeutic target; tool

DESCRIPTION (provided by applicant): Activation of the mitochondrial permeability transition pore (PTP) clearly plays a key role in some of the most wide-spread and therapeutically challenging human diseases. Our studies have established that the PTP operates in two modes 1) transiently, whereby the PTP acts as a mitochondrial Ca2+ release channel or 2) persistently, which ultimately results in cell death and disease. Although well characterized on a functional level, we know remarkably little about the molecules that form the PTP or how it is regulated. We urgently need answers to basic questions concerning what proteins actually form the PTP channel and what modulates the opening of the PTP in vivo. As a result, our goal is to identify the molecules that contribute to the structure and regulation of the PTP in both normal and disease states. This information is critical if we are to be able to effectively identify and/or deign valuable therapeutics targeting the transition of the PTP from normal to pathological. Here, we will use biochemical and genetic tools to identify structural components of the PTP and how PTP activity can be dynamically regulated in vivo. The specific objectives of this application are based in the synergy possible through the unique combination of novel approaches available in the Forte and Bernardi laboratories. The specific objectives of this application are: Aim 1 - Test role of OMM proteins in the regulation of PTP activity: While the PTP is primarily an IMM event, it has long been appreciated that proteins in the OMM should prominently regulate PTP activity. We will initially focus on a specific OMM protein, Tspo, whose role in PTP regulation has been strongly suggested. Our studies here will allow us to gain a deeper understanding of how cytosolic elements can impact PTP activity. Aim 2 - Identify key structural components of the PTP: Despite our increasing appreciation of its fundamental role in normal and pathological cellular responses, the molecules that form the PTP have remained a mystery. Here, we will use information in available mitochondrial proteomes to identify proteins forming the pore of the PTP. It is our expectation that the identification of any single component forming the PTP will supply us with the missing "hook", providing the opportunity to identify additional components. Aim 3 - Mitochondrial p66ShcA and ROS activation of the PTP: It is clear that pore open-closed transitions can be regulated at many levels and, by extension; misregulation of these upstream pathways can lead to persistent, pathological activation of the PTP. The goal of this aim is to investigate the hypothesis that ROS generated through the action of p66ShcA (p66) functions upstream of the activation of the PTP in conditions of excess oxidative stress. We anticipate on the completion of this aim to have clear understanding of the role of one novel upstream activator of PTP activity. These studies will set the stage for future interrogation aimed at extending our understanding of mitochondria and PTP activity in physiological and pathological settings. Clearly, these outcomes will be fundamental to developing novel therapeutic strategies specifically targeting the pore in the many disease processes in which the PTP has been clearly implicated. PUBLIC HEALTH RELEVANCE: The mitochondrial permeability transition pore has been studied for over 50 years and has been implicated, for example, in ischemia-reperfusion injury of the heart and brain, muscular dystrophy caused by collagen VI deficiency, and in the axonal damage occurring during MS among many other pathological conditions. Since little is known of the molecular composition of the PTP, our goals in this application are to use the pharmacological, biochemical and genetic tools we have established for the unbiased identification of proteins involved in the formation of the PTP and to use a variety of in vitro an in vivo tests to confirm their roles, either as core components of the pore itself, or regulators o pore activity. Since the PTP is of direct relevance to variety of human pathological conditions, we anticipate that the rigorous and careful identification of proteins forming or regulating the formation of the PTP will increase our ability to define therapies targeting these proteins as treatments for a wide variety of human diseases.

描述（由申请人提供）：线粒体通透性转换孔（PTP）的激活显然在一些最广泛和治疗上最具挑战性的人类疾病中起着关键作用。我们的研究已经确定PTP以两种模式运作：1）瞬时性，PTP充当线粒体Ca 2+释放通道; 2）持续性，最终导致细胞死亡和疾病。虽然在功能水平上得到了很好的表征，但我们对形成PTP的分子及其调节方式知之甚少。我们迫切需要答案的基本问题，即哪些蛋白质实际上形成PTP通道和什么调节PTP在体内的开放。因此，我们的目标是识别在正常和疾病状态下对PTP的结构和调节做出贡献的分子。如果我们要能够有效地识别和/或设计有价值的治疗方法，靶向PTP从正常到病理的转变，则该信息是至关重要的。在这里，我们将使用生物化学和遗传学的工具，以确定PTP的结构组成部分，以及如何PTP活动可以在体内动态调节。本申请的具体目标是基于通过Forte和Bernardi实验室提供的新方法的独特组合可能产生的协同作用。本申请的具体目的是：目的1 -测试OMM蛋白在PTP活性调节中的作用：虽然PTP主要是IMM事件，但长期以来人们认识到OMM中的蛋白应显著调节PTP活性。我们将首先关注一个特定的OMM蛋白，Tspo，其在PTP调节中的作用已被强烈建议。我们在这里的研究将使我们能够更深入地了解胞质元素如何影响PTP活性。目标2 -确定PTP的关键结构成分：尽管我们越来越认识到它在正常和病理细胞反应中的基本作用，但形成PTP的分子仍然是一个谜。在这里，我们将使用现有的线粒体蛋白质组中的信息来识别形成PTP孔的蛋白质。我们期望，对形成PTP的任何单个组件的识别将为我们提供缺失的“钩”，从而提供识别其他组件的机会。目的3 -PTP的线粒体p66 ShcA和ROS活化：很明显，孔开闭转换可以在许多水平上调节，并且通过扩展;这些上游途径的误调节可以导致PTP的持续病理活化。本研究的目的是研究在过度氧化应激条件下，通过p66 ShcA（p66）作用产生的ROS在PTP激活上游发挥作用的假设。我们期望在完成这一目标，有一个新的PTP活性的上游激活剂的作用有清楚的了解。这些研究将为未来的审讯奠定基础，旨在扩大我们对线粒体和PTP活性在生理和病理环境中的理解。显然，这些结果将是根本的发展新的治疗策略，特别是针对孔在许多疾病的过程中，PTP已明确牵连。 公共卫生关系：线粒体渗透性转换孔已经被研究了超过50年，并且已经涉及例如心脏和脑的缺血-再灌注损伤、由胶原VI缺乏引起的肌营养不良症、以及在MS期间发生的轴突损伤以及许多其他病理状况。由于对PTP的分子组成知之甚少，我们在本申请中的目标是使用我们已经建立的药理学、生物化学和遗传学工具来无偏地鉴定参与PTP形成的蛋白质，并使用各种体外和体内试验来确认它们的作用，无论是作为孔本身的核心组分，还是孔活性的调节剂。由于PTP与各种人类病理状况直接相关，我们预期严格和仔细鉴定形成或调节PTP形成的蛋白质将提高我们将靶向这些蛋白质的疗法定义为多种人类疾病的治疗的能力。