Phospholipids and Mitochondrial Function

磷脂和线粒体功能

• 批准号: 7873382
• 负责人: WILLIAM DOWHAN
• 金额: $ 20.42万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-03 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7873382
• 关键词: 3-Dimensional; 3-Methylglutaconic aciduria type 2; Aging; Apoptosis; Binding; Biochemical; Biochemical Genetics; Biological Models; Cardiolipins; Cell Nucleus; Cell physiology; Communication; Complex; Coupled; Cryoelectron Microscopy; Cytochrome bc1 Complex; Cytoplasm; Data; Detergents; Disease; Docking; Electron Microscopy; Electron Transport; Electron Transport Complex III; Energy Metabolism; Eukaryotic Cell; Functional disorder; Funding; Future; Genes; Genetic; Genetic Translation; Glues; Glycogen Branching Enzyme; Green Fluorescent Proteins; Heart failure; High Pressure Liquid Chromatography; Individual; Inner mitochondrial membrane; Ischemia; Light; Lipids; Location; Mammalian Cell; Messenger RNA; Mitochondria; Mitochondrial Diseases; Mitochondrial Proteins; Molecular; Molecular Genetics; Myocardial dysfunction; Negative Staining; Nuclear; Pathway interactions; Phase; Phenotype; Phosphatidyl glycerol; Phosphatidylglycerols; Phospholipids; Physiological; Play; Polyacrylamide Gel Electrophoresis; Property; Regulation; Reperfusion Therapy; Reporting; Research; Resolution; Role; Saccharomyces cerevisiae; Sampling; Signal Pathway; Site; Staging; Stress; Structure; System; Translation Initiation; Translations; Untranslated Regions; Yeasts; analog; base; biological adaptation to stress; complex IV; cytochrome c oxidase; dodecyl maltoside; essential phospholipids; mutant; novel; oligomycin sensitivity-conferring protein; particle; reconstitution; research study; response; stoichiometry; three dimensional structure

DESCRIPTION (provided by applicant): Cardiolipin (CL) is an essential phospholipid for normal mitochondrial energy metabolism so that physiological and pathological perturbations in its levels result in alterations in the structure, function and assembly of mitochondria. CL is an integral part of most of the components of the mitochondrial energy transducing system. Myocardial dysfunction and apoptosis are associated with abnormal CL levels in Barth Syndrome, aging, ischemia/reperfusion, and heart failure. Saccharomyces cerevisiae mutants (crd1?) lacking CL, but containing its precursor phosphatidylglycerol (PG), or mutants (pgs1?) lacking both PG and CL display similar phenotypes to mammalian cells with reduced PG and CL levels or the inability to make PG and CL, respectively. Therefore, yeast is an excellent model system to study their role of these lipids in mitochondrial function. Studies of yeast pgs1? and crd1? mutants led to two findings that are the basis for the proposed studies. (1) Translation repressors that bind 5' of the translation initiation site on mRNA for subunit 4 (Cox4p) of the mitochondrial electron transport chain component cytochrome c oxidase (Complex IV) are specifically activated or synthesized in response to altered mitochondrial phospholipid composition in pgs1? mutants. Studies are proposed to genetically and biochemically define the components and mechanism of this novel mitochondrial stress response pathway. (2) Complex IV and Complex III (cytochrome bc1) form a III2IV2 supercomplex that kinetically behaves as a single unit "respirasome", which was established by using a crd1? mutant to be specifically dependent on CL. Experiments are proposed to determine the phospholipid stoichiometry in the individual Complexes III and IV and in the III2IV2 supercomplex and the location of CL within the latter required for formation and stabilization of the supercomplex. A 3-D structure of the III2IV2 supercomplex has been determined by negative stain-electron microscopy and single particle analysis, which will be refined using cryoelectron microscopy to establish how Complex III and IV are organized in the supercomplex. Phospholipid analysis and structural data will be integrated to establish how CL is involved in "gluing" together the supercomplex components. Defining the roles PG and CL play in normal mitochondrial function will shed light on the molecular basis for cellular dysfunction in physiological and pathological states where these lipids are reduced.

说明(申请人提供)：心磷脂(CL)是正常线粒体能量代谢所必需的磷脂，其水平的生理和病理扰动会导致线粒体结构、功能和组装的改变。CL是线粒体能量转导系统中大多数组成部分的组成部分。心肌功能障碍和细胞凋亡与Barth综合征、衰老、缺血/再灌注和心力衰竭的CL水平异常有关。酿酒酵母突变株(crd1？)缺乏CL，但含有其前体磷脂酰甘油(PG)或突变体(Pgs1？)缺乏PG和CL的细胞表现出与哺乳动物细胞相似的表型，分别是PG和CL水平降低或不能产生PG和CL。因此，酵母是研究这些脂质在线粒体功能中的作用的一个很好的模型系统。酵母菌Pgs1？而crd1呢？突变导致了两个发现，这两个发现是拟议研究的基础。(1)线粒体电子传递链亚单位4(Cox4p)的翻译起始点的5‘端结合翻译抑制物细胞色素c氧化酶(Complex IV)是针对Pgs1中线粒体磷脂成分的改变而特异性激活或合成的。变种人。从遗传学和生物化学的角度对线粒体应激反应途径的组成和机制进行了研究。(2)络合物IV和络合物III(细胞色素Bc1)形成一个III2IV2超络合物，动力学上表现为一个单一的单位“呼吸体”，它是用crd1？突变体对CL具有特定的依赖性。建议通过实验来确定单独的络合物III和IV以及III2IV2超络合物中的磷脂化学计量比，以及超络合物形成和稳定所需的CL在后者中的位置。通过负染色电子显微镜和单粒子分析确定了III2IV2超络合物的三维结构，并将用低温电子显微镜对其进行精炼，以确定络合物III和IV在超络合物中的组织方式。磷脂分析和结构数据将被整合在一起，以确定CL是如何参与将超复杂成分粘合在一起的。明确PG和CL在正常线粒体功能中所起的作用将有助于阐明在这些脂质减少的生理和病理状态下细胞功能障碍的分子基础。