The molecular mechanisms of mitochondrial behavior.

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 保护细胞免于死亡，因此为治疗方法保护心脏提供了一个有效的靶点 从疾病的进展或心肌梗死事件。线粒体融合是由下列成员介导的 动力蛋白相关蛋白(DRP)家族，大的GTP酶，通过它们的自组装和 水解GTP，控制膜重塑事件。在哺乳动物中，Mfn1和Mfn2的功能取代了 单一外膜DRP在简单生物体中普遍表达，但相对表达 每种物质的水平都以组织依赖的方式变化。尽管Mfn1和Mfn2都在线粒体中发挥作用 融合，它们并不是完全多余的，尽管功能差异的性质尚不清楚。我们的 数据表明，Mfn1-Mfn2异型反式复合体的融合效率明显高于 要么是同型复合体。由此，我们预测Mfn1和Mfn2具有不同的分子性质和 这两个外膜DRP在哺乳动物中的功能意义是提供了一个相对简单的 它们通过各自和相对的表达水平调控融合机制。此外，数据表明， 存在特定的调控机制，因为我们已经证明，可溶性bax仅刺激Mfn2同型 复合体。为了了解两个外膜融合蛋白的功能意义以及它们之间的相互作用 调控它们活性的调节机制，我们建议表征它们的生化特性 每个DRP包括GTP结合、GTP水解和复杂组装。这些生化方法 共同决定了为什么只有Mfn1、Mfn2和Mfn1/Mfn2介导的机制基础 融合效率是不同的，可能会指出它们在细胞中受到明显调控的原因和方式。至 探索促凋亡的Bcl2蛋白Bax对Mfn2的哪些特性的调节，我们将测试 BAX对Mfn2的生化特性的影响，在心脏中具有特殊意义，其中Mfn2是 主要表达融合DRP。我们将进一步研究Bax对融合的调控。 相互作用结构域的鉴定及其在体外和细胞内的功能意义。BAX也 在被凋亡信号激活后对线粒体融合产生负面影响，我们将调查 融合的抑制是直接的或通过外膜通透性介导的。就像用 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 和线粒体的运动性，但这一点的意义相对来说还没有被探索。我们认为线粒体 微管连接和短程运动都是线粒体融合的重要调节因子。我们会 确定线粒体运动和拴系在调节线粒体融合和 用体外线粒体研究在这些过程中需要微核细胞的分子基础 融合动力分析和互补的生化方法。

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.