Systems Analysis of Mammalian Mitochondrial DNA Segregation

 描述(由申请人提供)：线粒体DNA(MtDNA)的忠实遗传对细胞呼吸是必不可少的，因此对大多数真核生物来说也是必不可少的。然而，在哺乳动物细胞中控制线粒体DNA维持和分离的途径还不是很清楚。这项工作的目标是了解哺乳动物线粒体DNA调控的分子机制。一旦知道了这一信息，就有可能确定影响人类健康的线粒体DNA单倍型分离的因素。具体地说，这项建议将解决线粒体DNA-蛋白质复合体，称为类核，在哺乳动物细胞中的行为。在目标1中，将使用超分辨率和共聚焦显微镜观察类核复合体，并结合遗传方法来耗尽和修饰复杂组分，以研究类核分离的细胞内基础，并测试候选效应蛋白的参与。目标2将阐述类核是如何组织或与细胞信号通路整合的。这将通过系统地绘制哺乳动物线粒体DNA类核复合体的物理和遗传相互作用图来实现。这些实验将提供对线粒体DNA可遗传单位的物理和功能组织的洞察，并有效地绘制出类核结构的完整的“相互作用体”。

 DESCRIPTION (provided by applicant): Faithful inheritance of mitochondrial DNA (mtDNA) is essential for cellular respiration and thus for most eukaryotic life. However, pathways that control mtDNA maintenance and segregation in mammalian cells are not well understood. The goal of this work is to understand the molecular mechanisms underlying the regulation of mammalian mtDNA. Once this information is known, it will be possible to identify factors that contribute to the segregation of mtDNA haplotypes that impact human health. Specifically, this proposal will address how mtDNA-protein complexes, termed nucleoids, behave in mammalian cells. In Aim 1, super-resolution and confocal microscopy of nucleoid complexes coupled with genetic approaches to deplete and modify complex components will be employed to investigate the intracellular basis of nucleoid segregation and test the involvement of candidate effector proteins. Aim 2 will address how the nucleoid is organized or integrated with cellular signaling pathways. This will be accomplished by systematically mapping physical and genetic interactions of the mammalian mtDNA nucleoid complex. These experiments will provide insight into the physical and functional organization of the heritable unit of mtDNA and effectively map the integrated 'interactome' of the nucleoid structure.