Mitochondrial Fusion and Division

描述(申请人提供)：线粒体是一个动态的膜结合细胞器，经历融合和分裂。这些相互对立的事件以协调的方式发生，它们之间的平衡是细胞器大小、数量和形状的关键决定因素。线粒体融合和分裂是由保守的动力蛋白相关GTP酶介导的，包括MFn(哺乳动物)/Fzo1p(酵母)融合和Drp1(哺乳动物)/Dnm1p(酵母)分裂。线粒体融合和分裂的异常与许多神经退行性疾病有关，如Charcot-Marie-Tooth神经病、显性视神经萎缩、阿尔茨海默病、亨廷顿病和帕金森病。这些疾病中的许多都会影响有丝分裂后神经元，这些神经元沿着其长轴突和分支树突含有线粒体。要了解这些疾病的发病机制，需要更深入地了解介导和协调线粒体融合和分裂的分子机制以及这些事件的生理功能。拟议的研究将揭示线粒体如何融合(目标1)，线粒体融合和线粒体分裂是如何协调的(目标2)，以及线粒体分裂如何控制有丝分裂后神经元中的线粒体分布(目标3)。为了研究Aim 1中线粒体融合的分子机制，我们纯化了线粒体融合所需的两种酵母蛋白-Fzo1p GTP酶和Fzo1p结合蛋白Ugo1p。利用这些蛋白质，我们已经开发了GTP结合、GTP水解和GTP依赖的膜融合的分析方法。这些新的检测方法将使我们能够剖析Fzo1p GTP酶和Ugo1p在线粒体融合中的功能。在目标2中，我们将确定线粒体融合和分裂是如何协调的。我们已经证明，在Drp1缺失的小鼠胚胎成纤维细胞中，Drp1的缺失降低了Mfn1和Mfn2的水平。我们将确定Drp1水平的变化如何转化为MFN水平的调节和线粒体融合。在目标3中，我们将通过使用Cre-loxP系统从小鼠有丝分裂后神经元中删除Drp1来确定线粒体分裂在神经元中的生理作用。初步数据显示，Drp1基因缺失会导致线粒体分布改变和神经变性。我们将确定线粒体分裂如何控制有丝分裂后神经元中细胞器的形态和分布。这项拟议研究的成功完成将提供对线粒体融合、平衡线粒体融合和分裂的协调机制以及线粒体分裂在神经元中的生理作用的机械性见解。 公共卫生相关性：线粒体融合和分裂的异常与许多神经疾病有关。为了更好地了解这些疾病的发病机制，我们将研究线粒体融合和分裂的分子机制和生理功能。

DESCRIPTION (provided by applicant): The mitochondrion is a dynamic membrane-bound organelle that undergoes fusion and division. The balance between these opposing events, which occur in a coordinated manner, is a key determinant of organelle size, number, and shape. Mitochondrial fusion and division are mediated by conserved dynamin-related GTPases including Mfn (mammals)/Fzo1p (yeast) for fusion and Drp1 (mammals)/Dnm1p (yeast) for division. Abnormalities in mitochondrial fusion and division are associated with many neurodegenerative diseases such as Charcot-Marie-Tooth neuropathy, dominant optic atrophy, Alzheimer's disease, Huntington's disease, and Parkinson's disease. Many of these diseases affect postmitotic neurons, which contain mitochondria along their long axons and branched dendrites. Understanding the pathogenesis of these diseases requires a deeper knowledge of the molecular mechanisms that mediate and coordinate mitochondrial fusion and division as well as the physiological functions of these events. The proposed research will uncover how mitochondria fuse (Aim 1), how mitochondrial fusion and mitochondrial division are coordinated (Aim 2), and how mitochondrial division controls mitochondrial distribution in postmitotic neurons (Aim 3). To study the molecular mechanisms underlying mitochondrial fusion in Aim 1, we have purified and biochemically characterized two yeast proteins that are required for mitochondrial fusion- Fzo1p GTPase and the Fzo1p- binding protein Ugo1p. Using these proteins, we have developed assays for GTP binding, GTP hydrolysis, and GTP-dependent membrane fusion. These novel assays will allow us to dissect the functions of Fzo1p GTPase and Ugo1p in mitochondrial fusion. In Aim 2, we will determine how mitochondrial fusion and division are coordinated. We have shown that the loss of Drp1 reduces Mfn1 and Mfn2 levels in Drp1-null mouse embryonic fibroblasts. We will determine how changes in Drp1 levels are translated into regulation of Mfn levels and mitochondrial fusion. In Aim 3, we will determine the physiological roles of mitochondrial division in neurons by deleting Drp1 from postmitotic neurons using the Cre-loxP system in mice. Preliminary data show that Drp1 loss induces alterations in mitochondrial distribution and neurodegeneration. We will determine how mitochondrial division controls organelle morphology and distribution in postmitotic neurons. Successful completion of the proposed studies will provide mechanistic insights into mitochondrial fusion, the coordination mechanism that balances mitochondrial fusion and division, and the physiological role of mitochondrial division in neurons. PUBLIC HEALTH RELEVANCE: Abnormalities in mitochondrial fusion and division are associated with many neurological disorders. To gain a better understanding of the pathogenesis of these diseases, we will investigate the molecular mechanisms and physiological functions of mitochondrial fusion and division.