Mutagenesis, tumorigenesis and human DNA polymerase epsilon

 描述（由申请方提供）：正常人体细胞具有遗传稳定性，自发突变率极低。相比之下，癌细胞通常在遗传上不稳定，并且在其基因组中积累了数千至数十万个突变。虽然已知DNA聚合酶校对有助于复制的准确性，但关于校对功能的丧失如何导致癌症的几个关键问题仍然没有答案：（1）校对功能障碍导致基因组不稳定性的机制是什么？(2)聚合酶校对的缺陷在多大程度上导致了癌症？以及（3）环境因素如金属暴露在多大程度上影响聚合酶依赖性肿瘤的发展？在这里，我们提供的证据表明，人类DNA聚合酶（Pol）ε（一种主要的复制型DNA聚合酶）中的癌症相关突变会损害其校对活性，导致独特类型的诱变增加，并在金属暴露期间提供生存优势。我们假设这些效应对肿瘤发生有直接影响。 该项目的主要目标是验证我们的中心假设，即Pol ε的体细胞突变通过几种非排他性机制为肿瘤发展提供了选择性优势，包括（1）肿瘤抑制基因中失活无义突变的积累;（2）对氧化DNA损伤剂的抗性，包括重金属暴露。这个假设是基于我们的初步数据。具体而言，本项目将1）建立致癌Pol ε突变体的动力学基础 等位基因; 2）定义Pol ε核酸外切酶结构域突变（EDM）产生其独特突变特征的机制;和3）确定Pol ε突变促进肿瘤发展的机制。 拟议的研究是创新的，因为多学科的方法，结合体外和体内研究，以表征癌症相关的Pol ε突变对基因组稳定性的影响。对复制叉缺陷如何影响基因组改变的新见解也是创新的。这一贡献是重要的，因为它将提供新的和详细的见解的生化机制如何复制的DNA聚合酶通常防止收购的复杂多样性的突变中发现的癌症基因组，以及提供见解的基本机制的DNA复制。这些知识将加深我们对癌症发展的理解，并最终为未来旨在调节DNA聚合酶活性的研究提供信息，以实现新的癌症治疗策略的目标。

 DESCRIPTION (provided by applicant): Normal somatic human cells are genetically stable and have a very low spontaneous mutation rate. In contrast, cancer cells are typically genetically unstable and accumulate thousands to hundreds of thousands of mutations in their genome. While it is known that DNA polymerase proofreading contributes to the accuracy of replication, several critical questions regarding how the loss of the proofreading function contributes to cancer remain unanswered: (1) What are the mechanisms through which proofreading dysfunction contributes to genome instability? (2) What extent do defects in polymerase proofreading play in driving cancer? and (3) To what extent do environmental factors like metal exposure influence polymerase-dependent tumor development? Here we provide evidence that cancer-associated mutations in human DNA polymerase (Pol) ε, a major replicative DNA polymerase, impair its proofreading activity, cause an increase in a unique type of mutagenesis and provide a survival advantage during metal exposure. We hypothesize that these effects have a direct influence on tumorigenesis. The main goal of this project is to test our central hypothesis that somatic mutations in Pol ε provide a selective advantage to tumor development through several, non-exclusive mechanisms, including (1) the accumulation of inactivating nonsense mutations in tumor suppressor genes; (2) resistance to oxidizing DNA damaging agents, including heavy metal exposure. This hypothesis is based on our preliminary data. Specifically, this project will 1) Establish a kinetic basis for cancer-causing Pol ε mutant alleles; 2) Define the mechanisms through which Pol ε exonuclease domain mutations (EDMs) generate their unique mutational signatures; and 3) Determine the mechanisms through which mutations in Pol ε contribute to tumor development. The proposed research is innovative due to the multidisciplinary approach that combines in vitro and in vivo studies to characterize the effects of cancer-associated Pol ε mutations on genome stability. The novel insights into how defects at the replication fork can influence genomic alterations are also innovative. This contribution is significant because it will provide new and detailed insights into the biochemical mechanisms of how replicative DNA polymerases normally prevent the acquisition of the complex diversity of mutations found in cancer genomes, as well as provide insights into the fundamental mechanisms of DNA replication. This knowledge will deepen our understanding of cancer development and can ultimately serve to inform future studies designed to modulate DNA polymerase activities toward the goal of novel cancer therapeutic strategies.