Organization, Expression, and Inheritance of Yeast mtDNA

描述（由申请人提供）： 拟议研究的总体目标是了解遗传油线粒体DNA（mtDNA）的机制，使用芽殖酵母，酿酒酵母，作为模式生物。我们的研究将结合联合收割机遗传学、生物化学、分子生物学和细胞生物学的方法来表征营养生长过程中有助于mtDNA忠实传递的各种成分。 为此，我们提出了以下具体目标：1）探讨线粒体DNA类核的结构和组织。目的是了解类核蛋白如何与mtDNA结合，以及它们如何促进mtDNA的组织和遗传; 2）阐明mtDNA分离装置的组成部分。我们将使用各种生物化学和细胞生物学方法来研究mtDNA类核蛋白和某些突变形式的类核蛋白，Ilv 5 p，它的行为根据所有测试标准像mtDNA类核蛋白，如何与线粒体内膜和候选蛋白质，可能是mtDNA分离装置的组成部分相互作用; 3）阐明RTG-逆行反应途径在线粒体DNA稳定性和遗传中的作用。我们已经发现，这一途径是密切参与线粒体DNA遗传通过其控制的“代谢”类核蛋白。我们将探讨这条逆行途径是如何控制线粒体DNA的稳定性和遗传性的，以及如何绕过对线粒体DNA主要包装蛋白Abf 2 p的要求; 4）建立线粒体DNA类核分裂的规律，以及热休克蛋白60（Hsp 60）、活性ori序列和转录在类核分裂中的作用。这些目标来自我们的发现，即类核分裂是一个受调控的过程。我们建议开发异步和同步类核分裂系统。这些研究将与区分活性和非活性ori元件的顺式作用元件的分析相结合。总之，这些研究对理解人类线粒体疾病有直接影响。

DESCRIPTION (provided by applicant): The overall objective of the proposed research is to understand the mechanism of inheritance oil mitochondrial DMA (mtDNA) using the budding yeast, Saccharomyces cerevisiae, as a model organism. Our studies will combine genetic, biochemical, molecular and cell biological approaches to characterize the various components that contribute to the faithful transmission of mtDNA during vegetative growth. To these ends, we propose the following specific aims: 1) to probe the structure and organization of mtDNA nucleoids. The goal is to understand how the nucleoid proteins are associated with mtDNA, and how they contribute to mtDNA organization and inheritance; 2) to elucidate components of the mtDNA segregation apparatus. We will use a variety of biochemical and cell biological methods to examine how mtDNA nucleoids and certain mutant forms of the nucleoid protein, Ilv5p, which behaves according to all criteria tested like mtDNA nucleoids, interact with the inner mitochondrial .membrane and with candidate proteins that are likely to be components of the mtDNA segregation apparatus; 3) to elucidate the role of the RTG-retrograde response pathway in mtDNA stability and inheritance. We have found that this pathway is intimately involved in mtDNA inheritance through its control of 'metabolic' nucleoid proteins. We will evaluate how the retrograde pathway controls the stability and inheritance of mtDNA, and circumvents the requirement for the major mtDNA packaging protein, Abf2p; 4) to establish the rules of mtDNA nucleoid division and the roles of Hsp60, active ori sequences and transcription of mtDNA in nucleoid division. These objectives follow from our discovery that nucleoid division is a regulated process. We propose to develop both asynchronous and synchronous nucleoid division systems. These studies will be coupled with the analysis of cis-acting elements that distinguish active from inactive ori elements. Altogether, these studies have direct implications for understanding mitochondrial diseases in humans.