Translesion DNA polymerase kappa activity in gliomas

描述（由申请人提供）：转录DNA合成聚合酶（TLS pols）对于绕过和耐受DNA损伤至关重要。TLS pols的错误调节发生在许多类型的癌症中，包括胶质源性的恶性脑肿瘤。人类DNA聚合酶κ（hpol κ）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复制的精确影响仍然未知。本文概述的实验将检查芳香烃受体（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。中心目标包括研究促进胶质瘤细胞中hpol κ过表达的机制，确定这种错误调节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，并研究通过靶向抑制脑肿瘤中的TLS pols来改善胶质瘤治疗的潜在方法。目前的提案将解决重要的问题，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配体影响hpol κ表达和/或活性吗？3)阻断胶质瘤内源性AhR信号传导如何影响复制应激？4)胶质瘤中TLS活性的调节能增强抗癌治疗吗？5)hpol κ和基因组看守Werner综合征蛋白（WRN）之间的蛋白质-蛋白质相互作用是否限制了神经胶质瘤细胞中DNA损伤的突变旁路？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应激和TLS活性的模型，为我们了解神经胶质瘤中的基因组维持和诱变提供信息，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，这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.