Institute of Radiation Biology, Helmholtz Centre Munich - German Research Centre for Environmental Health, Ingolstaedter Landstrasse 1, D-85764 Neuherberg, Germany.
Nucleic Acids Res. 2010 Apr;38(6):1853-65. doi: 10.1093/nar/gkp1175. Epub 2009 Dec 29.
The Mre11/Rad50/Xrs2 (MRX) complex in Saccharomyces cerevisiae has well-characterized functions in DNA double-strand break processing, checkpoint activation, telomere length maintenance and meiosis. In this study, we demonstrate an involvement of the complex in the base excision repair (BER) pathway. We studied the repair of methyl-methanesulfonate-induced heat-labile sites in chromosomal DNA in vivo and the in vitro BER capacity for the repair of uracil- and 8-oxoG-containing oligonucleotides in MRX-deficient cells. Both approaches show a clear BER deficiency for the xrs2 mutant as compared to wildtype cells. The in vitro analyses revealed that both subpathways, long-patch and short-patch BER, are affected and that all components of the MRX complex are similarly important for the new function in BER. The investigation of the epistatic relationship of XRS2 to other BER genes suggests a role of the MRX complex downstream of the AP-lyases Ntg1 and Ntg2. Analysis of individual steps in BER showed that base recognition and strand incision are not affected by the MRX complex. Reduced gap-filling activity and the missing effect of aphidicoline treatment, an inhibitor for polymerases, on the BER efficiency indicate an involvement of the MRX complex in providing efficient polymerase activity.
酿酒酵母中的 Mre11/Rad50/Xrs2（MRX）复合物在 DNA 双链断裂处理、检查点激活、端粒长度维持和减数分裂中具有明确的功能。在这项研究中，我们证明了该复合物参与碱基切除修复（BER）途径。我们研究了体内甲基甲磺酸酯诱导的热不稳定位点在染色质 DNA 中的修复，以及在 MRX 缺陷细胞中修复含有尿嘧啶和 8-氧鸟嘌呤的寡核苷酸的体外 BER 能力。与野生型细胞相比，这两种方法都清楚地显示出 xrs2 突变体的 BER 缺陷。体外分析表明，长补丁和短补丁 BER 这两个亚途径都受到影响，MRX 复合物的所有成分对于 BER 的新功能同样重要。对 XRS2 与其他 BER 基因的上位性关系的研究表明，MRX 复合物在 AP 裂合酶 Ntg1 和 Ntg2 下游发挥作用。对 BER 中各个步骤的分析表明，碱基识别和链切割不受 MRX 复合物的影响。缺口填充活性降低，以及聚酶抑制剂 aphidicoline 处理对 BER 效率的缺失效应表明，MRX 复合物参与提供有效的聚酶活性。
