Roles of Human DNA Polymearse Epsilon in Mutagenesis and Genome Stability

项目总结(见说明)： 环境侮辱会导致DNA碱基损伤和DNA链断裂，如果不加以修复，这些结果可能会破坏基因组的稳定。为了避免基因组的不稳定性和癌症，DNA聚合酶在准确复制基因组和修复受损的DNA方面发挥着核心作用。DNA聚合酶E(Pol E)不仅在复制和修复中起重要作用，而且在DNA聚合酶中是独一无二的，它通过检查点功能参与协调复制进程和有丝分裂。 Pol e在复制和修复过程中扮演的重要但尚未明确的角色，使得对其保真度的研究对于理解人类细胞如何能够避免DNA损伤的后果至关重要。如果不修复DNA损伤，DNA损伤可能会导致突变和癌症等疾病。鉴于它的重要性，对人类Pol e保真度的缺乏与关于酵母酶保真度的信息量的增长形成了鲜明的对比。确定DNA聚合酶对基因组稳定性的贡献的一个重要工具是鉴定和表征DNA聚合酶的突变衍生物，这些突变会降低DNA合成的整体保真度。该项目建议识别和表征人类Pol e的突变等位基因 通过降低复制保真度，改变病变旁路，抑制体内Pol e检查点的激活 效率和忠诚度，或这些活动的某种组合。此外，这些等位基因将被用来探测Pol e在体内复制和修复中的功能。 酵母和人类Pol e都已在参与复制的多蛋白复合体中被发现， 转录和重组，并已观察到在体内与活性区域共定位 复制。这些相互作用仍然没有得到很好的描述，也没有被很好地理解，特别是对人类酶。该项目建议通过使用以下方法来确定POL？蛋白质相互作用网络 重组Pol-e探查Pol-e相互作用伙伴及其功能意义 互动。

PROJECT SUMMARY (See instructions): Environmental insults can lead to DNA base damages and DNA strand breaks, outcomes that have the potential to destabilize the genome if left unrepaired. In order to avoid genome instability and cancer, DNA polymerases play a central role in accurately replicating the genome and repairing damaged DNA. DNA Polymerase e (Pol E) specifically plays important roles not only in replication and repair, but is unique among DNA polymerases in its involvement in coordinating replication progression and mitosis through its checkpoint function. The important, yet not precisely defined roles played by Pol e in replication and repair make the study of its fidelity crucial to understanding how human cells are able to avoid the consequences of DNA damage that, if left unrepaired, can lead to mutagenesis and diseases like cancer. Given its importance, the lack of knowledge of the fidelity of human Pol e stands in sharp contrast to the growing amount of information on the fidelity of the yeast enzyme. One important tool to determine the contribution of a DNA polymerase to genome stability is to identify and characterize mutant derivatives of DNA polymerases that decrease overall DNA synthesis fidelity. This project proposes to identify and characterize mutant alleles of human Pol e that suppress Pol e checkpoint activation in vivo through decreased replication fidelity, altered lesion bypass efficiency and fidelity, or some combination of these activities. In addition, these alleles will be used to probe for Pol e function in replication and repair in vivo. Both yeast and human Pol e have been identified in multi-protein complexes involved in replication, transcription and recombination, and have been observed to colocalize in vivo with regions of active replication. These interactions remain poorly characterized and not well understood, especially for the human enzyme. This project proposes to determine the Pol ¿ protein interaction network by using recombinant Pol e to probe for Pol e interaction partners and characterize the functional significance of these interactions.