Mechanisms of damaged DNA replication in eukaryotes

描述（由申请人提供）：我们研究的长期目标是在热力学、动力学和结构水平上了解真核生物中受损DNA的复制。模板链中的DNA损伤阻断了经典DNA聚合酶的复制。因此，细胞具有多种非经典DNA聚合酶，其可以取代停滞在DNA损伤位点的经典聚合酶，并且可以通过损伤进行复制。最近的动力学研究和结构研究提供了大量的见解，这些非经典聚合酶如何不同于经典聚合酶，以及它们如何能够适应DNA损伤。然而，目前尚不清楚非经典聚合酶如何被募集到DNA损伤位点，停滞的经典聚合酶如何从DNA损伤位点被置换，以及复制辅助因子如何促进与非经典聚合酶的DNA损伤相反的核苷酸掺入。为了解决这些问题，我们提出了以下三个具体目标的研究：（1）以确定其他蛋白质因子对核苷酸掺入的影响相反的DNA损伤的非经典聚合酶，（2）以确定机制的非经典聚合酶招聘在translesion合成，和（3）以确定机制的经典聚合酶位移在translesion合成。这些研究将提供一个清晰的理解，确切地说，非经典的聚合酶如何取代经典的聚合酶在DNA损伤的网站，以及其他蛋白质因子如何有助于复制受损的DNA。此外，这些研究将有助于我们了解突变和癌症的起源，并将提供对其预防的见解。

DESCRIPTION (provided by applicant): The long-term goal of our research is to understand the replication of damaged DNA in eukaryotes at the thermodynamic, kinetic, and structural levels. DNA damage in the template strand blocks replication by classical DNA polymerases. Consequently cells possess a variety of non-classical DNA polymerases that can replace the classical polymerase stalled at sites of DNA damage and can replicate through the damage. Recent kinetic studies and structural studies have provided substantial insights into how these non-classical polymerases differ from classical polymerases and how they are able to accommodate DNA damage. It remains unclear, however, how non-classical polymerases are recruited to sites of DNA damage, how stalled classical polymerases are displaced from sites of DNA damage, and how replication accessory factors promote nucleotide incorporation opposite DNA damage by non-classical polymerases. To address these issues, we propose studies with the following three specific aims: (1) to determine the effect of other protein factors on nucleotide incorporation opposite DNA damage by non-classical polymerases, (2) to determine the mechanism of non-classical polymerase recruitment during translesion synthesis, and (3) to determine the mechanism of classical polymerase displacement during translesion synthesis. These studies will provide a clear understanding of exactly how non-classical polymerases replace classical polymerases at sites of DNA damage and how other protein factors contribute to the replication of damaged DNA. Furthermore, these studies will contribute to our understanding of the origins of mutations and cancers and will provide insight into their prevention.