Dissecting the mechanism of increased genomic instability in aging yeast

描述（由申请人提供）：年龄是最大的致癌物，然而随着人类年龄的增长，癌症发病率急剧增加的机制尚不清楚。为了更好地理解这一现象，我们在酵母酿酒酵母中开发了一个系统，该系统概括了癌症的标志之一——基因组不稳定性——作为细胞年龄的功能。具体来说，我们通过监测酵母在复制衰老过程中不同染色体的杂合性损失（LOH）来研究衰老与基因组不稳定性之间的关系。我们发现，衰老的酵母母细胞在其后代中经历了LOH增加近100倍的转变。最近，我们对这一现象有了更好的理解，并提出了一个有效的假设，将与年龄相关的LOH分解为一系列相互关联的事件。它始于mtDNA的丢失或损坏。这导致线粒体内膜电位的降低，进而导致铁硫簇（ISC）生物合成和/或ISC组装成维持基因组完整性所需的蛋白质的缺陷。这些基因组完整性蛋白的功能降低导致LOH水平升高。这个假设可以作为一个工作模型来形成我们的问题和方法。我们现在建议使用s.s erevisiae中可用的强大资源和工具集来测试这一假设中的步骤。作为该方法的一部分，我们开发了一种诱导遗传系统，即母亲富集程序（MEP），该系统首次允许方便地分离复制老化的酿酒酵母细胞进行生化、细胞生物学和遗传分析。这将使我们能够进一步了解衰老过程中最早的事件，最终导致与年龄相关的LOH。因为在这个过程中，每一个与年龄相关的LOH都是保守的，我们相信我们在这个计划中所学到的东西最终将被转化为识别人类中同样受衰老影响并导致年龄相关肿瘤发生的候选基因和过程。

DESCRIPTION (provided by applicant): Age is the greatest carcinogen, yet the mechanism by which the incidence of cancer dramatically increases as humans age is unclear. In an effort to better understand this phenomenon, we developed a system in the yeast Saccharomyces cerevisiae that recapitulates one of the hallmarks of cancer - genomic instability - as a function of cellular age. Specifically, we examined the relationship between aging and genomic instability by monitoring loss of heterozygosity (LOH) on different chromosomes during replicative aging in yeast. We discovered that aging yeast mother cells underwent a switch to a nearly 100-fold increase in LOH in their progeny. Recently, we gained a better understanding of this phenomenon and have developed a working hypothesis that breaks the age-associated LOH into a series of connected events. It begins with a loss of, or damage to, the mtDNA. This leads to a reduction in the inner mitochondrial membrane potential (??), which in turn leads to a defect in iron-sulfur cluster (ISC) biosynthesis and/or assembly of the ISCs into proteins required for maintenance of genome integrity. The reduced function of these genome integrity proteins then leads to increased levels of LOH. This hypothesis serves as a working model to formulate our questions and approaches. We now propose to test the steps in this hypothesis using the powerful collection of resources and tools available in S. cerevisiae. As part of this approach, we developed an inducible genetic system, the Mother Enrichment Program (MEP), which permits for the first time the facile isolation of replicatively aged S. cerevisiae cells for biochemical, cell biological and genetic analysis. This will permit us to gain further understanding of the earliest events in aging that ultimately lead to age-associated LOH. Because each of the processes in this progression to age-associated LOH are conserved, we believe that what we learn in the proposed plan will ultimately be translatable to identifying candidate genes and processes in humans that are similarly affected by aging and lead to age-related oncogenesis. PUBLIC HEALTH RELEVANCE: There is a striking link between increasing age and the incidence of cancer in humans. Though the simple organism baker's yeast does not get cancer, we have found that older yeast cells exhibit a dramatic increase in one of the hallmarks of cancer: genomic instability. In this proposal we will use yeast as a model system to identify and understand the factors responsible for this instability in order to gain insight into how age impacts the incidence of oncogenesis.