Deng Changchun, Brown James A, You Dongqing, Brown J Martin
Department of Radiation Oncology, Stanford University School of Medicine, California 94305-5152, USA.
Genetics. 2005 Jun;170(2):591-600. doi: 10.1534/genetics.104.028795. Epub 2005 Apr 16.
Topoisomerase I plays a vital role in relieving tension on DNA strands generated during replication. However if trapped by camptothecin or other DNA damage, topoisomerase protein complexes may stall replication forks producing DNA double-strand breaks (DSBs). Previous work has demonstrated that two structure-specific nucleases, Rad1 and Mus81, protect cells from camptothecin toxicity. In this study, we used a yeast deletion pool to identify genes that are important for growth in the presence of camptothecin. In addition to genes involved in DSB repair and recombination, we identified four genes with known or implicated nuclease activity, SLX1, SLX4, SAE2, and RAD27, that were also important for protection against camptothecin. Genetic analysis revealed that the flap endonucleases Slx4 and Sae2 represent new pathways parallel to Tdp1, Rad1, and Mus81 that protect cells from camptothecin toxicity. We show further that the function of Sae2 is likely due to its interaction with the endonuclease Mre11 and that the latter acts on an independent branch to repair camptothecin-induced damage. These results suggest that Mre11 (with Sae2) and Slx4 represent two new structure-specific endonucleases that protect cells from trapped topoisomerase by removing topoisomerase-DNA adducts.
拓扑异构酶I在缓解复制过程中DNA链上产生的张力方面起着至关重要的作用。然而，如果被喜树碱或其他DNA损伤所捕获，拓扑异构酶蛋白复合物可能会使复制叉停滞，从而产生DNA双链断裂（DSB）。先前的研究表明，两种结构特异性核酸酶Rad1和Mus81可保护细胞免受喜树碱毒性的影响。在本研究中，我们使用酵母缺失文库来鉴定在喜树碱存在下对生长至关重要的基因。除了参与DSB修复和重组的基因外，我们还鉴定出四个具有已知或潜在核酸酶活性的基因，即SLX1、SLX4、SAE2和RAD27，它们对抵御喜树碱也很重要。遗传分析表明，瓣状核酸内切酶Slx4和Sae2代表了与Tdp1、Rad1和Mus81平行的新途径，可保护细胞免受喜树碱毒性的影响。我们进一步表明，Sae2的功能可能归因于其与核酸内切酶Mre11的相互作用，并且后者作用于一个独立的分支来修复喜树碱诱导的损伤。这些结果表明，Mre11（与Sae2一起）和Slx4代表两种新的结构特异性核酸内切酶，它们通过去除拓扑异构酶-DNA加合物来保护细胞免受被困拓扑异构酶的影响。
