The molecular mechanisms of mitochondrial behavior.

线粒体行为的分子机制。

• 批准号: 8875045
• 负责人: Suzanne C Hoppins
• 金额: $ 23.99万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-16 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8875045
• 关键词: Address; Apoptosis; Apoptotic; Bax protein; Behavior; Biochemical; Biological Assay; Cardiac; Cell Death; Cell physiology; Cells; Characteristics; Chimeric Proteins; Chronic; Complex; Coupled; Data; Deletion Mutation; Disease Progression; Dynamin; Embryo; Event; Foundations; GTP Binding; Grant; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Heart; Heart Diseases; Hydrolysis; In Vitro; Knowledge; Light; Link; Mammals; Maps; Mediating; Membrane; Membrane Fusion; Microtubules; Mitochondria; Molecular; Movement; Myocardial Infarction; Nature; Neurodegenerative Disorders; Organelles; Organism; Pathway interactions; Physiological; Play; Process; Property; Protein Family; Proteins; Regulation; Relative (related person); Role; Shapes; Signal Pathway; Signal Transduction; Testing; Therapeutic; Time; Tissues; Translating; abstracting; base; cell motility; chemical genetics; drug development; fusion gene; genetic approach; in vitro Assay; insight; member; novel; protein function; research study; therapeutic development

Project Summary/Abstract: Mitochondrial fusion regulates the shape, distribution and function of the organelle and plays critical roles in protection from cell death and therefore offers a valid target for theurapeutic approaches to protect the heart from progression of disease or myocardial infarction events. Mitochondrial fusion is mediated by members of the dynamin related protein (DRP) family, large GTPases that, through their ability to self-assemble and hydrolyze GTP, control membrane remodeling events. In mammals, MFN1 and MFN2 function in place of a single outer membrane DRP in simple organisms, both are expressed ubiquitously but the relative expression level of each varies in a tissue dependent manner. Although MFN1 and MFN2 both function in mitochondrial fusion, they are not completely redundant, although the nature of the functional differences are not known. Our data indicate that the MFN1-MFN2 heterotypic trans complex is significantly more efficacious for fusion than either homotypic complex. From this, we predict that MFN1 and MFN2 have distinct molecular properties and that the functional significance of the two outer membrane DRPs in mammals is to provide a relatively simple regulatory fusion mechanism via their respective and relative expression levels. Further, data suggest that specific regulatory mechanisms exist, as we have shown that soluble Bax stimulates only MFN2 homotypic complexes. In order to understand the functional significance of two outer membrane fusion proteins and the regulatory mechanisms that govern their activity, we propose to characterize the biochemical properties of each DRP including GTP binding, GTP hydrolysis and complex assembly. These biochemical approaches together will determine the mechanistic basis for why MFN1 only, MFN2 only and MFN1/MFN2 mediated fusion efficiencies are distinct and will likely point to why and how they are distinctly regulated in cells. To explore what properties of MFN2 are modulated by the pro-apoptotic Bcl2 protein Bax, we will test the effect of Bax on the biochemical properties of MFN2, of particular significance in the heart where MFN2 is the predominantly expressed fusion DRP. We will further characterize the regulation of fusion by Bax through identification of interaction domains and exploring their functional significance in vitro and in cells. Bax also negatively effects mitochondrial fusion following activation by apoptotic signals and we will investigate whether the inhibition of fusion is direct or mediated through outer membrane permeabilization. As observed with the apoptotic pathway and mitochondrial dynamics, an interdependence also exists between mitochondrial fusion and mitochondrial motility, but the significance of this is relatively unexplored. We propose that mitochondrial tethering to microtubules and short range motility are both important regulators of mitochondrial fusion. We will determine the role that mitochondrial movement and tethering play in regulating mitochondrial fusion and investigate the molecular basis for the requirement of MFNs in these processes using an in vitro mitochondrial fusion-motility assay and complimentary biochemical approaches.

项目摘要/摘要： 线粒体融合调节细胞器的形状，分布和功能，并在 防止细胞死亡，因此为保护心脏提供了有效的目标 来自疾病或心肌梗塞事件的进展。线粒体融合由 与动力蛋白相关的蛋白质（DRP）家族，大的GTPases，通过它们的自我组装能力和 水解GTP，控制膜重塑事件。在哺乳动物中，MFN1和MFN2功能代替A 简单生物中的单个外膜DRP，两者都普遍表达，但相对表达 每种水平都以组织依赖性方式变化。尽管MFN1和MFN2都在线粒体中起作用 融合，它们不是完全冗余的，尽管功能差异的性质尚不清楚。我们的 数据表明，MFN1-MFN2异型式复合物比融合更有效 要么同型复合物。由此，我们预测MFN1和MFN2具有不同的分子特性，并且 哺乳动物中两个外膜DRP的功能意义是提供一个相对简单的 通过各自和相对表达水平的调节融合机制。此外，数据表明 存在特定的调节机制，因为我们已经表明可溶性BAX仅刺激MFN2同型 复合物。为了了解两种外膜融合蛋白的功能意义和 管理其活动的监管机制，我们建议表征 每个DRP，包括GTP结合，GTP水解和复合组装。这些生化方法 共同确定为什么仅MFN1，仅MFN2和MFN1/MFN2介导的机械基础 融合效率是不同的，可能会指出为什么在细胞中明显调节它们的原因和方式。到 探索MFN2的哪些特性是通过促凋亡的Bcl2蛋白Bax调节的，我们将测试 在MFN2的生化特性上的Bax，在MFN2为心脏中特别重要的是 主要表达融合DRP。我们将进一步表征通过Bax对融合的调节 鉴定相互作用域并探索其在体外和细胞中的功能意义。也是Bax 凋亡信号激活后的线粒体融合产生负面影响，我们将研究是否研究 融合的抑制是通过外膜透化直接或介导的。如与 凋亡途径和线粒体动力学，线粒体融合之间也存在相互依赖性 和线粒体运动性，但其意义相对尚未探索。我们提出了线粒体 将微管束缚和短距离运动都是线粒体融合的重要调节剂。我们将 确定线粒体运动和绑扎在调节线粒体融合和 使用体外线粒体研究这些过程中MFN的分子基础 融合动力测定和免费生化方法。