Molecular Dissection of the Permeability Transition Pore

渗透率转变孔的分子解剖

• 批准号: 7214064
• 负责人: MICHAEL A FORTE
• 金额: $ 29.58万
• 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 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7214064
• 关键词: Adenine Nucleotides; Affinity; Apoptosis; Apoptotic; Attention; BCL-2 Protein; BCL2 gene; Bioenergetics; Biological; Biological Assay; Cell Death; Cell Membrane Permeability; Cell Survival; Cells; Charge; Chemicals; Complex; Cyclosporine; Data; Development; Diagnostic; Disease; Dissection; Energy Metabolism; Event; Family; Family member; Goals; Homeostasis; Human; Immunosuppressive Agents; In Vitro; Inner mitochondrial membrane; Knock-out; Laboratories; Lead; Life; Maintenance; Mediator of activation protein; Membrane; Membrane Potentials; Metabolism; Mitochondria; Modeling; Modification; Molecular; Multiprotein Complexes; Mus; Mutant Strains Mice; Nerve Degeneration; Outer Mitochondrial Membrane; Participant; Pathologic Processes; Permeability; Play; Process; Production; Protein Isoforms; Proteins; Proton Pump; Protons; Reagent; Reperfusion Injury; Respiration; Ro 68-3400; Role; Rupture; Site; Standards of Weights and Measures; Structure; System; Testing; Tissues; VDAC1 gene; Yeasts; analog; base; cell growth regulation; cyclophilin D; cytochrome c; experience; genetic regulatory protein; human disease; in vivo; inhibitor/antagonist; mitochondrial dysfunction; mitochondrial permeability transition pore; molecular mass; novel; novel strategies; programs; pyridine nucleotide; research study; respiratory; solute; tool

Mitochondria playa pivotal role in cell survival and tissue development by virtue of their role in energy metabolism, regulation of cellular Ca2 + homeostasis and apoptosis. Given this multifactorial role, they regulate cellular Ca2+ metabolism and bioenergetics function as an integrated s~stem. In terms of normal physiology, this integration is reflected in mitochondrion's high capacity to store Ca2 +, which may protect cells like neurons against transient elevation in intracellular Ca2 + during periods of hyperactivity. Mitochondrial Ca2 + homeostasismust be tightly regulated and is based in a series of specific uptake and release systems. Yet, in vitro themitochondrial inner membrane (IMM) can easily undergo a permeability increase to solutes with molecularmasses of about 1,500 Da or lower. This permeability change, called the permeability transition (PT), isregulated by the opening of a membrane pore, the mitochondrial permeability transition pore (PTP). The PTPis voltage-dependent, cyclosporin A (CsA)-sensitive, high-conductance channel of the inner mitochondrialmembrane; pore open-closed transitions are highly regulated by multiple effectors that likely converge on asmaller set of regulatory sites. The PTP has long been implicated as a target for mitochondrial dysfunction invivo, particularly in the context of specific human pathological events. These suspicions have been confirmedby examination of mice in which the expression of mitochondrial CyPD (a key regulator of PTP action and thetarget of CsA) has been eliminated. These studies have confirmed a critical role for the PTP in models of ischemia/reperfusion injury both in the heart and the brain, models of muscular dystrophy, in the axonaldamage occurring during MS, and Alzheimer's disease. However, despite detailed functional characterizationover the last 30 years, none of the candidate pore components in traditional models has withstood critical andunambiguous genetic tests. In this light, the PBR remains the only biochemically identified component in traditional molecular models of the PTP that has not been subjected to thorough genetic testing. Consequently, the overall goal of this application is to use biochemical and genetic tools to critically test the role of the peripheral benzodiazepine receptor (PBR) in PTP function using a variety of in vitro and in vivo tests that we have developed to confirm its role. either as core components or regulators of the PTP. Our studies are based in mice that we have now successfully generated in which the wild-type Tspo gene has been replaced with a modified Tspo gene containing /oxP sites. Using these mice, our plan is to test PTP function in mitochondria, cells and tissues lacking the PBR and thereby rigorously evaluate the role of the PBR in PTP function. Importantly. since mice have now been successfully generated containing a modified Tspo gene containing loxP sites. it is reasonable that we will be able to complete these aims in the two years of funding provided.

线粒体Playa Playa关键在细胞存活和组织发育中的作用，凭借其在 能量代谢，细胞Ca2 +稳态的调节和凋亡。给定这个多因素 角色，它们调节细胞Ca2+代谢和生物能力作为综合S〜茎的作用。在 正常生理学的术语，这种整合反映在线粒体存储Ca2的高能力中 +，可以保护像神经元这样的细胞 在多动症周期内对细胞内Ca2 +的短暂升高。线粒体Ca2 + 稳态受到严格的调节，并基于一系列特定的摄取和释放系统。 然而，在体外主题软骨内膜（IMM）很容易经历渗透性增加到 分子量约为1,500 da或更低的溶质。这种渗透性变化，称为 通过膜孔的打开，线粒体的开口，渗透率转变（PT） 渗透性过渡孔（PTP）。 PTPIS电压依赖性，环孢菌素A（CSA）敏感， 线粒体内膜内部的高导通道；孔闭合过渡是 高度受到多种效应子的调节，这些效应可能会收敛于Asmaller的监管部位。这 长期以来，PTP一直被认为是线粒体功能障碍的目标，特别是在 特定人类病理事件的背景。这些怀疑已得到证实 线粒体CYPD表达的小鼠（PTP动作和thetarget的关键调节剂 CSA）已被消除。这些研究已经证实了PTP在模型中的关键作用 心脏和大脑中的缺血/再灌注损伤，肌肉营养不良的模型， 在MS和阿尔茨海默氏病期间发生的轴突症。但是，尽管功能详细 在过去的30年中，表征传统模型中没有一个候选孔成分 经受了批判性和无歧义的基因检测。从这个角度看，PBR仍然是唯一的 生化鉴定的成分 未经彻底基因检测的PTP的传​​统分子模型。 因此，该应用程序的总体目标是使用生化和遗传工具进行批判性 使用多种IN测试外围苯二氮卓受体（PBR）在PTP功能中的作用 我们已经开发出来确认其作用的体外和体内测试。作为核心组件或 PTP的调节器。我们的研究基于我们现在成功产生的小鼠 野生型TSPO基因已被替换为 修饰的TSPO基因含有 /OXP位点。使用这些鼠标，我们的计划是测试PTP功能 线粒体，细胞和组织缺乏PBR，从而严格评估PBR在 PTP功能。重要的是。由于现在已经成功地生成了一个修改后的小鼠 含有LOXP位点的TSPO基因。合理的是，我们将能够在两者中完成这些目标 提供了多年的资金。