Rinne M L, He Y, Pachkowski B F, Nakamura J, Kelley M R
Department of Biochemistry and Molecular Biology, Indiana University School of Medicine Indianapolis, IN 46202, USA.
Nucleic Acids Res. 2005 May 19;33(9):2859-67. doi: 10.1093/nar/gki601. Print 2005.
Previous studies indicate that overexpression of N-methylpurine DNA glycosylase (MPG) dramatically sensitizes cells to alkylating agent-induced cytotoxicity. We recently demonstrated that this sensitivity is preceded by an increased production of AP sites and strand breaks, confirming that overexpression of MPG disrupts normal base excision repair and causes cell death through overproduction of toxic repair intermediates. Here we establish through site-directed mutagenesis that MPG-induced sensitivity to alkylation is dependent on enzyme glycosylase activity. However, in contrast to the sensitivity seen to heterogeneous alkylating agents, MPG overexpression generates no cellular sensitivity to MeOSO2(CH2)2-lexitropsin, an alkylator which exclusively induces 3-meA lesions. Indeed, MPG overexpression has been shown to increase the toxicity of alkylating agents that produce 7-meG adducts, and here we demonstrate that MPG-overexpressing cells have dramatically increased removal of 7-meG from their DNA. These data suggest that the mechanism of MPG-induced cytotoxicity involves the conversion of non-toxic 7-meG lesions into highly toxic repair intermediates. This study establishes a mechanism by which a benign DNA modification can be made toxic through the overexpression of an otherwise well-tolerated gene product, and the application of this principle could lead to improved chemotherapeutic strategies that reduce the peripheral toxicity of alkylating agents.
先前的研究表明，N-甲基嘌呤DNA糖基化酶（MPG）的过表达会使细胞对烷化剂诱导的细胞毒性显著敏感。我们最近证明，这种敏感性之前会伴随AP位点和链断裂的产生增加，证实MPG的过表达会破坏正常的碱基切除修复，并通过过量产生有毒的修复中间体导致细胞死亡。在这里，我们通过定点诱变确定，MPG诱导的烷基化敏感性取决于酶的糖基化酶活性。然而，与对异质烷化剂的敏感性不同，MPG过表达不会使细胞对MeOSO2(CH2)2-lexitropsin产生敏感性，MeOSO2(CH2)2-lexitropsin是一种专门诱导3-甲基腺嘌呤（3-meA）损伤的烷化剂。事实上，MPG过表达已被证明会增加产生7-甲基鸟嘌呤（7-meG）加合物的烷化剂的毒性，并且在这里我们证明，过表达MPG的细胞从其DNA中去除7-meG的能力显著增强。这些数据表明，MPG诱导细胞毒性的机制涉及将无毒的7-meG损伤转化为高毒性的修复中间体。这项研究确立了一种机制，通过该机制，一种良性的DNA修饰可以通过过量表达一种原本耐受性良好的基因产物而变得有毒，并且这一原理的应用可能会导致改进的化疗策略，从而降低烷化剂的外周毒性。
