Translesion Synthesis DNA Polymerases and Genome Instability

描述（由申请人提供）：由外源性基因毒物自发或诱导的突变是癌症的根本原因。几乎所有基因毒物诱导的突变都是由于DNA损伤和特殊的翻译合成（TLS） DNA聚合酶对受损DNA的复制，其准确性低于正常的复制DNA聚合酶。TLS聚合酶在复制未受损DNA时也非常容易出错，因此构成了基因组不稳定的持续来源，需要加以控制以避免疾病。虽然TLS聚合酶募集到DNA损伤位点的机制已经被很好地理解，但限制它们参与正常复制的机制却不太清楚。PI的实验室发现DNA聚合酶？（Pol?）在未受损DNA的复制过程中受到多种阻碍复制进程的因素的促进，包括正常复制机制中的缺陷，叉在自然障碍处停滞以及治疗性复制抑制剂的治疗。本提案旨在定义调节Pol？在体内DNA损伤和生理条件下。在具体目标1中，我们将确定dNTP池的检查点依赖提升在形成Pol的错误签名中的作用。在具体目标2中，我们将确定Pol？DNA合成受复制动力学和分叉不对称的调控。目标1和目标2的研究将利用酵母模型系统，目的是利用在酵母中获得的数据进一步推进我们对人类细胞诱变机制的理解。在Specific Aim 3中，我们将描述人类细胞中对复制缺陷的诱变反应的机制。拟议的工作将导致更好地理解在正常细胞中运作的诱变过程，以及由环境基因毒物或治疗干预引起的诱变过程。从长远来看，学会操纵这些过程将有助于减少癌症发病率，延缓进展和改善治疗效果。

DESCRIPTION (provided by applicant): Mutations occurring spontaneously or induced by exogenous genotoxicants are a root cause of cancer. Nearly all genotoxicant-induced mutations result from DNA damage and replication of the damaged DNA by specialized translesion synthesis (TLS) DNA polymerases that are less accurate than normal replicative DNA polymerases. TLS polymerases are also highly error-prone when copying undamaged DNA, thus constituting a persistent source of genomic instability that needs to be controlled to avoid disease. While the mechanisms of TLS polymerase recruitment to the sites of DNA damage are understood relatively well, the mechanisms that restrict their participation in the normal replication are much less clear. The PI's laboratory has discovered that the participation of DNA polymerase ? (Pol?) in the copying of undamaged DNA is promoted by a variety factors that impede the progression of the replication, including defects in the normal replication machinery, fork stalling at natural impediments and treatment with therapeutic replication inhibitors. This proposal seeks to define the global mechanisms that regulate the extent of error-prone synthesis by Pol? in vivo in DNA damaging and physiological conditions. In Specific Aim 1, we will determine the role of checkpoint dependent elevation of dNTP pools in shaping the error signature of Pol?. In Specific Aim 2, we will determine how the contribution of Pol? to DNA synthesis is regulated by the replication dynamics and fork asymmetry. The yeast Saccharomyces cerevisiae model system will be utilized in the studies proposed in Aims 1 and 2, with the goal of using the data obtained in yeast to further advance our understanding of the mechanisms of mutagenesis in human cells. In Specific Aim 3, we will characterize the mechanism of the mutagenic response to replication defects in human cells. The proposed work will lead to a better understanding of the mutagenic processes that operate in normal cells, as well as those induced by environmental genotoxicants or therapeutic interventions. In the long run, learning to manipulate these processes will help reduce cancer incidence, delay progression and improve therapy outcome.