Translesion DNA polymerase kappa activity in gliomas

描述（由申请人提供）：翻译DNA合成聚合酶（TLS pol）对于绕过和耐受DNA损伤至关重要。TLS极点的错误调控发生在许多类型的癌症中，包括源自神经胶质的恶性脑肿瘤。人类DNA聚合酶kappa （hpol κ）最近被证明是胶质瘤患者预后不良的独立预后指标。然而，导致胶质瘤中hpol κ上调的机制以及对DNA复制的确切影响尚不清楚。本文概述的实验将研究芳烃受体（AhR）的促肿瘤激活是否导致胶质瘤中hpol κ的组成性上调，然后研究这些致命肿瘤中与hpol κ活性相关的机制特征。主要目标包括研究促进胶质瘤细胞中hpol κ过表达的机制，确定这种错误调控对复制应激反应的影响，研究改变hpol κ活性的蛋白质-蛋白质相互作用，以及通过靶向抑制脑肿瘤中的TLS pol来改善胶质瘤治疗的潜在方法。目前的提案将解决一些重要的问题，例如：1)AhR通路是否调节胶质瘤中的hpol κ ？2)外源性或内源性AhR配体是否影响hpol κ的表达和/或活性？3)阻断胶质瘤内源性ahr信号如何影响复制应激？4)胶质瘤中TLS活性的调节是否能增强抗癌治疗？5) hpol κ和基因组看守Werner's综合征蛋白（WRN）之间的蛋白-蛋白相互作用是否限制了胶质瘤细胞中DNA损伤的诱变旁路？实验策略利用体外细胞培养、诱变分析、结构-功能分析、小分子抑制剂研究和蛋白质组学来确定hpol κ在胶质瘤中的作用。该应用程序的长期目标是产生复制压力和TLS活性的模型，以告知我们对胶质瘤中基因组维持和突变的理解，同时也为我们如何更好地治疗患有这种疾病的患者提供见解。从这些研究中得出的结论将大大改善我们对恶性脑肿瘤过程的基本定义，恶性脑肿瘤是一个主要的公共卫生问题。

DESCRIPTION (provided by applicant): Translesion DNA synthesis polymerases (TLS pols) are vital to bypass and tolerance of DNA damage. The mis-regulation of TLS pols occurs in many types of cancer, including malignant brain tumors of glial origin. Human DNA polymerase kappa (hpol κ) was recently shown to be an independent prognostic indicator of poor outcome in glioma patients. However, the mechanism(s) leading to up-regulation of hpol κ in gliomas and the precise effects upon DNA replication remain unknown. The experiments outlined herein will examine whether tumor-promoting activation of the aryl hydrocarbon receptor (AhR) leads to constitutive up-regulation of hpol κ in gliomas and then investigate mechanistic features associated with hpol κ activity in these deadly tumors. The central objectives include investigating mechanisms that promote hpol κ over-expression in glioma cells, determining the effect of this mis-regulation upon replication stress responses, studying protein-protein interactions that alter hpol κ activity and examining potential ways of improving glioma treatment through targeted inhibition of TLS pols in brain tumors. The current proposal will address important questions, such as: 1) Does the AhR pathway regulate hpol κ in gliomas? 2) Do exogenous or endogenous AhR ligands affect hpol κ expression and/or activity? 3) How does blocking endogenous AhR-signaling in gliomas affect replication stress? 4) Can modulation of TLS activity in gliomas potentiate anti-cancer treatments? 5) Do protein-protein interactions between hpol κ and the genomic caretaker Werner's syndrome protein (WRN) limit mutagenic bypass of DNA damage in glioma cells? The experimental strategies utilize in vitro cell culture, mutagenesis assays, structure-function analyses, small-molecule inhibitor studies and proteomics to ascertain the role of hpol κ in gliomas. The long-term goal of the application is to produce models for replication stress and TLS activity that inform our understanding of genomic maintenance and mutagenesis in gliomas, while also providing insights into how we might better treat patients suffering from this disease. The conclusions derived from these studies will dramatically improve our fundamental definition of processes that occur in malignant brain tumors, a major public health concern.