Trapped topoisomerase II initiates formation of de novo duplications via the nonhomologous end-joining pathway in yeast
Trapped topoisomerase II initiates formation of de novo duplications via the nonhomologous end-joining pathway in yeast
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DOI：
10.1073/pnas.2008721117
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发表时间：
2020-10-27
影响因子：
11.1
通讯作者：
Nitiss, John L.
中科院分区：
文献类型：
--
作者：
Stantial, Nicole;Rogojina, Anna;Nitiss, John L.
拓扑异构酶II （Top2）是分解姐妹染色单体之间的连环烷以及与双链DNA的过度或欠缠绕相关的超级线圈的重要酶。Top2通过在DNA中制造双链断裂（DSB）并通过断裂传递完整的双链来改变DNA拓扑结构。Top2同型二聚体的每个组成单体划伤一条DNA链，并与5'端形成共价磷酸酪氨酸键。化疗药物（如依托泊苷）稳定这种中间体可导致持久性和潜在毒性的DSB5。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等位基因的过表达与一个突变特征相关，其特征是依赖于DSB修复的非同源末端连接途径的DNA序列的重新复制。从DNA末端清除酶可以促进top2相关的复制，当作为寡核苷酸的一部分去除蛋白质时，top2相关的复制被抑制。用依托泊苷处理的TOP2细胞表现出相同的突变特征，过表达野生型蛋白的细胞也是如此。这些结果对基因组进化具有启示意义，并且与靶向Top2的化疗药物的临床使用有关。
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 DSB5. 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.