Topoisomerase II (Top2) is an essential enzyme that resolves catenanes between sister chromatids as well as supercoils associated with the over- or under-winding of duplex DNA. Top2 alters DNA topology by making a double-strand break (DSB) in DNA and passing an intact duplex through the break. Each component monomer of the Top2 homodimer nicks one of the DNA strands and forms a covalent phosphotyrosyl bond with the 5′ end. Stabilization of this intermediate by chemotherapeutic drugs such as etoposide leads to persistent and potentially toxic DSBs. We describe the isolation of a yeast top2 mutant (top2-F1025Y,R1128G) the product of which generates a stabilized cleavage intermediate in vitro. In yeast cells, overexpression of the top2-F1025Y,R1128G allele is associated with a mutation signature that is characterized by de novo duplications of DNA sequence that depend on the nonhomologous end-joining pathway of DSB repair. Top2-associated duplications are promoted by the clean removal of the enzyme from DNA ends and are suppressed when the protein is removed as part of an oligonucleotide. TOP2 cells treated with etoposide exhibit the same mutation signature, as do cells that overexpress the wild-type protein. These results have implications for genome evolution and are relevant to the clinical use of chemotherapeutic drugs that target Top2.

拓扑异构酶II（Top2）是一种必不可少的酶，它可以解决姐妹染色单体之间的链烷以及与双链DNA过度缠绕或缠绕不足相关的超螺旋。Top2通过在DNA中进行双链断裂（DSB），并使完整的双链体通过断裂来改变DNA拓扑。Top2同型二聚体的每个组成单体都在DNA链中形成一条切口，并与5'端形成共价磷酸酪氨酰键。通过化学疗法药物（如依托泊苷）稳定该中间体会导致持久性和潜在毒性的DSB。我们描述了酵母top2突变体（top2-F1025Y，R1128G）的分离，其产物在体外产生稳定的裂解中间体。在酵母细胞中，top2-F1025Y，R1128G的过表达等位基因与突变特征相关，其特征在于DNA序列从头重复，这取决于DSB修复的非同源末端连接途径。从DNA末端清除酶会促进与Top2相关的复制，当蛋白质作为寡核苷酸的一部分被清除时，Top2相关的复制会受到抑制。依托泊苷处理过的TOP2细胞与过表达野生型蛋白的细胞具有相同的突变特征。这些结果对基因组进化有影响，并且与靶向Top2的化学治疗药物的临床应用有关。