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）：线粒体通透性过渡孔（PTP）的激活在某些最广泛且具有治疗挑战性的人类疾病中显然起着关键作用。我们的研究已经确定，PTP以两种模式运行1）瞬时，PTP充当线粒体Ca2+释放通道或2）持久，最终导致细胞死亡和疾病。尽管在功能水平上表征良好，但我们对形成PTP的分子或调节方式的分子知之甚少。我们迫切需要解决有关哪种蛋白质实际构成PTP通道以及调节体内PTP开放的基本问题的答案。结果，我们的目标是确定在正常和疾病状态下有助于PTP的结构和调节的分子。如果我们能够有效地识别和/或取消针对PTP从正常情况过渡到病理的过渡的有价值的治疗剂，那么此信息至关重要。在这里，我们将使用生化和遗传工具来识别PTP的结构成分以及如何在体内动态调节PTP活性。该应用程序的具体目标是通过在Forte和Bernardi实验室中提供的新颖方法的独特组合来基于协同作用。该应用程序的具体目标是：目标1- OMM蛋白在PTP活性调节中的测试作用：虽然PTP主要是IMM事件，但长期以来一直认为，OMM中的蛋白质应显着调节PTP活性。我们最初将重点放在特定的OMM蛋白TSPO上，该蛋白在PTP调节中的作用得到了强烈建议。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。 AIM 2-确定PTP的关键结构成分：尽管我们对其在正常和病理细胞反应中的基本作用越来越多，但形成PTP的分子仍然是一个谜。在这里，我们将在可用的线粒体蛋白质组中使用信息来识别形成PTP孔的蛋白质。我们期望识别任何形成PTP的单个组件将为我们提供缺失的“钩子”，从而提供了识别其他组件的机会。 AIM 3-线粒体P66SHCA和PTP的ROS激活：很明显，可以在许多级别上调节孔隙开放式过渡，并扩展；这些上游途径的不正调会导致PTP的持续性病理激活。该目的的目的是研究通过在过量氧化应激条件下激活PTP上游的P66SHCA（P66）函数的作用产生的ROS。我们预计该目标的完成将清楚地了解一种新型PTP活动上游激活因子的作用。这些研究将为未来的审讯奠定阶段，旨在扩展我们对生理和病理环境中的线粒体和PTP活性的理解。显然，这些结果将是制定新型的治疗策略的基础，这些治疗策略是针对PTP明显涉及的许多疾病过程中毛孔的专门针对毛孔的。 公共卫生相关性：线粒体通透性过渡孔已经进行了50多年的研究，并被涉及，例如，在心脏和大脑的缺血 - 重新灌注损伤中，由胶原蛋白VI缺乏症引起的肌肉性营养不良，以及在许多其他病理条件下发生的轴突损伤引起的。由于对PTP的分子组成知之甚少，因此我们在本应用中的目标是使用我们确定的药理学，生化和遗传工具来无公正地鉴定参与PTP形成的蛋白质，并使用各种体外的体外体体内测试来确认其角色，以确认其核心组成的核心成分，或者是孔子的核心组成部分。由于PTP与人类病理状况的种类直接相关，因此我们预计，对形成或调节PTP形成的蛋白质的严格而仔细的鉴定将增加我们定义靶向这些蛋白质作为各种人类疾病的治疗方法的疗法的能力。