Phospholipids and Mitochondrial Function

磷脂和线粒体功能

基本信息

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
项目类别：
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

项目摘要

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.

描述（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，因此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 是线粒体能量转换系统大多数组件的组成部分。心肌功能障碍和细胞凋亡与巴斯综合征、衰老、缺血/再灌注和心力衰竭中异常的 CL 水平相关。缺乏 CL 但含有其前体磷脂酰甘油 (PG) 的酿酒酵母突变体 (crd1?) 或同时缺乏 PG 和 CL 的突变体 (pgs1?) 显示出与哺乳动物细胞相似的表型，但 PG 和 CL 水平降低或无法产生 PG 和 CL ，分别。因此，酵母是研究这些脂质在线粒体功能中的作用的绝佳模型系统。酵母pgs1的研究？和crd1？突变体导致了两项研究结果，这是拟议研究的基础。 (1) 结合线粒体电子传递链成分细胞色素 c 氧化酶（复合体 IV）亚基 4 (Cox4p) mRNA 翻译起始位点 5' 的翻译抑制子，响应 pgs1 中线粒体磷脂组成的改变而被特异性激活或合成？突变体。提出研究从遗传学和生物化学角度定义这种新型线粒体应激反应途径的组成部分和机制。 (2) 复合物 IV 和复合物 III (细胞色素 bc1) 形成 III2IV2 超级复合物，其动力学行为类似于单个单元“呼吸体”，该复合物是通过使用 crd1? 建立的。突变体特别依赖于 CL。建议进行实验以确定各个复合物 III 和 IV 以及 III2IV2 超级复合物中的磷脂化学计量，以及后者中形成和稳定超级复合物所需的 CL 位置。 III2IV2 超级复合物的 3D 结构已通过负染色电子显微镜和单粒子分析确定，并将使用冷冻电子显微镜对其进行细化，以确定复合物 III 和 IV 在超级复合物中的组织方式。磷脂分析和结构数据将被整合，以确定 CL 如何参与将超级复杂成分“粘合”在一起。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。