Mitochondria and Mutagenesis

描述(申请人提供)：线粒体功能障碍是癌症的标志之一。然而，人们对线粒体功能障碍如何导致癌症知之甚少。我们在这项建议中描述的研究表明，线粒体在保护细胞免受核基因组突变的影响方面发挥了新的作用，核基因组突变是人类癌症发生中的一个重要事件。以酿酒酵母为模式生物，分析了线粒体功能破坏对核基因组突变的影响。我们测量了刀豆素抗性菌落的频率，作为核基因组突变的指标。我们的数据表明，线粒体功能障碍导致(I)缺乏完整线粒体基因组(Rho？或p？)或缺失线粒体DNA(Rho-或p-)的突变菌株(线粒体遗传功能障碍，MGD)和(Ii)当野生型酵母中的氧化磷酸化被抗菌素A阻断时(线粒体代谢功能障碍，MMD)。MMD和MGD细胞的核突变频率分别高于未经处理的对照细胞和野生型细胞。MGD导致细胞内ROS水平降低。相反，MMD导致细胞内活性氧物种(ROS)水平增加。我们证明了MGD引起的核基因组突变依赖于Rev1、RevS或Rev7基因产物，所有这些都与复制后修复(PRR)有关。然而，MMD引起的核基因组突变不涉及Rev1、RevS或Rev7基因。此外，我们提供了Rtg2蛋白(逆行2)参与线粒体到细胞核的逆行反应途径保护细胞免受核基因组突变的证据。基于这些观察，我们假设线粒体保护并使用多种途径来保护核基因组免受突变的影响。我们提出了4个具体目标来检验所提出的假设。在所有情况下，我们都有为拟议的具体目标提供基础的初步数据。这些具体目标是：1)建立线粒体到核的逆行反应通路在线粒体功能障碍所致核基因组突变中的作用2)确定氧化应激反应受损是否导致核基因组突变3)复制后修复通路在线粒体功能障碍所致核基因组突变中的作用4)确定线粒体功能障碍所致核基因组突变的性质我们的研究将为深入了解线粒体功能障碍所致核基因组突变的分子遗传学机制提供帮助。

DESCRIPTION (provided by applicant): Mitochondrial dysfunction is one of the hallmarks of cancer. However, very little is known about how mitochondrial dysfunction leads to carcinogenesis. Our studies described in this proposal suggest a novel role for mitochondria in protecting cells from nuclear genome mutagenesis, an important event in human carcinogenesis. Using Saccharomyces cerevisiae as a model organism, we analyzed the consequences of disrupting mitochondrial function on mutagenesis of the nuclear genome. We measured the frequency of canavanine resistant colonies as an indicator of nuclear genome mutagenesis. Our data demonstrate that mitochondrial dysfunction leads to mutation in the nuclear genome (i) in mutant strains lacking the entire mitochondrial genome (rho¿ or p¿) or those with deleted mitochondrial DNA (rho- or p-) (mitochondrial genetic dysfunction, MGD) and (ii) when oxidative phosphorylation is blocked in wild type yeast by antimycin A (mitochondrial metabolic dysfunction, MMD). The nuclear mutation frequencies in both MMD and MGD cells were higher compared to untreated control and wild type cells respectively. MGD led to decreased intracellular levels of ROS. In contrast MMD led to increased intracellular levels of reactive oxygen species (ROS). We demonstrate that nuclear genome mutagenesis due to MGD is dependent on REV1, REVS or REV7 gene products, all implicated in post-replication repair (PRR). However, nuclear genome mutagenesis due to MMD does not involve REV1, REVS or REV7 genes. Furthermore, we provide evidence that Rtg2 protein (retrograde 2) involved in mitochondria-to-nucleus retrograde response pathway protect cells from nuclear genome mutagenesis. Based on these observations we hypothesize that mitochondria protect and employ multiple pathways to guard the nuclear genome against mutagenesis. We propose 4 specific aims to test the proposed hypothesis. In all cases we have preliminary data that provide the basis for the proposed specific aims. These specific aims are:1) Establish the role of mitochondria-to-nucleus retrograde response pathway in nuclear genome mutagenesis due to mitochondrial dysfunction 2) Determine whether compromised oxidative stress response leads to nuclear genome mutagenesis due to mitochondrial dysfunction 3) Determine the role of post-replication repair pathway in nuclear genome mutagenesis due to mitochondrial dysfunction 4) Determine the nature of nuclear genome mutagenesis due to mitochondrial dysfunction Our proposed study should provide insight into the molecular genetic mechanisms of nuclear genome mutagenesis due to mitochondrial dysfunction that is of fundamental significance to human cancer and other diseases.