Pol Epsilon Proofreading and Genetic Stability in Mice

描述（由申请人提供）：校对是DNA聚合酶保真度的主要监护人。真核生物有两种校对聚合酶，Pol 5和Pol e，它们是染色体DNA忠实复制所必需的。为了确定校对的生物学意义，我们制造了在这些必需聚合酶的核酸外切酶（exo）结构域中具有失活点突变的“敲入”小鼠。在这里，我们提出表征我们的新的Pol e exo~突变系，连同Pol 8 exo ~和错配修复（MMR）突变体，以描绘哺乳动物中控制忠实DNA合成和癌症避免的途径。本项目的总体目标是确定Pole校对的生物学功能。具体目标是：1）表征Pol e exo-小鼠的存活和癌症表型，2）表征Pol e exo-细胞和组织的突变子表型，3）确定哺乳动物Pol 6和Pol e校正的功能关系，以及4）确定Pol e校正和MMR在突变和癌症避免中的协同作用。总之，这些研究将揭示Pole校对对哺乳动物生物学，DNA复制保真度和癌症的重要性。这些研究为表征映射到Pol e校正域的人类多态性奠定了基础，并为开发利用复制错误灾难进行抗肿瘤治疗的新策略提供了工具。

DESCRIPTION (provided by applicant): Proofreading is the primary guardian of DNA polymerase fidelity. Eukaryotes have two proofreading polymerases, Pol 5 and Pol e that are required for faithful chromosomal DNA replication. To determine the biologic significance of proofreading, we made "knock-in" mice with inactivating point mutations in the exonuclease (exo) domains of these essential polymerases. Here we propose to characterize our new Pol e exo~ mutant line, together with Pol 8 exo" and mismatch repair (MMR) mutants, to delineate pathways that govern faithful DNA synthesis and cancer avoidance in mammals. The overall objective of this project is to determine the biologic functions of Pol e proofreading. The specific aims are: 1) characterize the survival and cancer phenotypes of Pol e exo" mice, 2) characterize the mutator phenotype of Pol e exo" cells and tissues, 3) determine the functional relationship of mammalian Pol 6 and Pol e proofreading, and 4) determine the cooperative roles of Pol e proofreading and MMR in mutation and cancer avoidance. Together, these studies will reveal the significance of Pol e proofreading to mammalian biology, DNA replication fidelity and cancer. These studies lay the groundwork for characterizing human polymorphisms that map to the proofreading domain of Pol e, and provide tools for developing novel strategies that exploit replication error-catastrophe for anti-tumor therapy.