Chan Katie, Houlbrook Sue, Zhang Qiu-Mei, Harrison Mark, Hickson Ian D, Dianov Grigory L
Radiation and Genome Stability Unit, Medical Research Council, Harwell, Oxfordshire OX11 0RD, UK.
Mutagenesis. 2007 May;22(3):183-8. doi: 10.1093/mutage/gel070. Epub 2007 Jan 31.
DNA polymerase beta (Pol beta) is important for the base excision repair (BER) pathway. Overexpression of Pol beta is frequently found in cancer cells and is thought to be associated with tumorigenesis. In this study, we examined BER fidelity in extracts derived from a human lymphoblastoid cell line that over expresses Pol beta compared to normal control cells. Using an in vitro mutagenesis assay, we found an increased rate of frameshift mutations arising during DNA repair in whole-cell extracts derived from the Pol beta-overexpressing cells. We demonstrate that the addition of excess Pol beta to a control cell extract enhances the mutagenic potential of the extract. Furthermore, using cell extracts and purified Pol beta, we demonstrate that the mechanism of frameshift formation involves slippage of Pol beta during the one-nucleotide gap-filling step of BER and that this slippage is fixed by strand-displacement synthesis stimulated by an excess of Pol beta.
DNA聚合酶β(Pol β)对碱基切除修复(BER)途径很重要。Pol β的过表达在癌细胞中经常被发现,并被认为与肿瘤发生有关。在本研究中,我们检测了与正常对照细胞相比,来自过表达Pol β的人淋巴母细胞系提取物中的BER保真度。使用体外诱变测定法,我们发现在来自过表达Pol β细胞的全细胞提取物中,DNA修复过程中产生的移码突变率增加。我们证明,向对照细胞提取物中添加过量的Pol β会增强提取物的诱变潜力。此外,使用细胞提取物和纯化的Pol β,我们证明移码形成的机制涉及BER的单核苷酸缺口填充步骤中Pol β的滑动,并且这种滑动通过过量的Pol β刺激的链置换合成得以固定。