Dissecting the mechanism of increased genomic instability in aging yeast

项目摘要 年龄是最大的致癌因素，然而，随着年龄的增长， 人类的年龄不清楚。为了更好地理解这种现象，我们在酵母中开发了一个系统， 酿酒酵母，概括了癌症的标志之一-基因组不稳定性-作为一个 细胞年龄的功能。具体来说，我们通过以下方式研究了衰老和基因组不稳定性之间的关系： 监测酵母复制老化过程中不同染色体上的杂合性丢失（洛）。我们 发现老化的酵母母细胞在其染色体上的洛缺失增加了近100倍。 后代 最近，我们对这一现象有了更好的了解，并提出了一个工作假设 将与年龄相关的洛缺失分解为一系列相关事件。它开始于一个损失，或损害， mtDNA。这导致线粒体内膜电位降低（），这反过来又导致 铁硫簇（ISC）生物合成和/或ISC组装成所需蛋白质的缺陷 保持基因组的完整性。这些基因组完整性蛋白的功能降低， 洛缺失水平升高。这一假设作为一个工作模型来制定我们的问题， 接近。 我们现在建议使用强大的资源和工具集合来测试这个假设中的步骤 在S.酿酒的。作为这种方法的一部分，我们开发了一种诱导型遗传系统， 富集计划（MEP），这使得第一次容易分离复制老化的S。 用于生物化学、细胞生物学和遗传分析的酿酒酵母细胞。这将使我们能够进一步获得 了解最终导致与年龄相关的洛缺失的衰老早期事件。因为每个 在这个发展过程中与年龄相关的洛缺失是保守的，我们相信，我们所了解的， 拟议的计划将最终转化为确定候选基因和人类的过程， 类似地受衰老影响并导致与年龄相关的肿瘤发生。

PROJECT SUMMARY 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.