DNA double-strand breaks induced by DNA topoisomerase IIα and reactive oxygen species lead to the integration of foreign DNA into the host genome

IF 2.4 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis Pub Date : 2026-01-01 Epub Date: 2025-12-04 DOI:10.1016/j.mrfmmm.2025.111924

Aya Kurosawa , Masumi Umehara , Noritaka Adachi

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引用次数: 0

Abstract

Foreign DNA integrates into the genome at low frequencies and random positions upon introduction into cells. This phenomenon, known as random integration, occurs when DNA double-strand breaks (DSBs) are repaired by non-homologous end-joining or polymerase theta-mediated end-joining, incorporating the foreign DNA into the genome. However, the mechanism underlying DSB generation in random integration remains unclear. In this study, we investigated the role of DNA topoisomerase II (Top2) and reactive oxygen species (ROS) in generating DSBs and examined their effects on random integration frequency. In Nalm-6 cells, the frequency of random integration increased following treatment with the Top2 inhibitor etoposide and hydrogen peroxide. Conversely, depletion of Top2α and cultivation under hypoxic conditions independently reduced the frequency of random integration, with their combination resulting in an even greater reduction. In gene targeting experiments at the HPRT locus, both Top2α depletion and hypoxic culture similarly reduced random integration. This suggests that DSBs generated by Top2α and ROS contribute to random integration and impede efficient gene targeting.

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DNA拓扑异构酶i α和活性氧诱导的DNA双链断裂导致外源DNA整合到宿主基因组中

外源DNA在进入细胞后以低频率和随机位置整合到基因组中。这种被称为随机整合的现象发生在DNA双链断裂（dsb）通过非同源末端连接或聚合酶介导的末端连接修复时，将外源DNA整合到基因组中。然而，DSB在随机整合中产生的机制尚不清楚。在这项研究中，我们研究了DNA拓扑异构酶II （Top2）和活性氧（ROS）在dsb生成中的作用，并研究了它们对随机整合频率的影响。在Nalm-6细胞中，用Top2抑制剂依托泊苷和过氧化氢处理后，随机整合的频率增加。相反，Top2α的消耗和缺氧条件下的培养单独降低了随机整合的频率，它们的结合导致更大的减少。在HPRT基因位点的基因靶向实验中，Top2α缺失和缺氧培养同样减少了随机整合。这表明由Top2α和ROS产生的dsb促进了随机整合，阻碍了有效的基因靶向。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis 生物-毒理学

CiteScore

4.90

自引率

0.00%

发文量

审稿时长

51 days

期刊介绍： Mutation Research (MR) provides a platform for publishing all aspects of DNA mutations and epimutations, from basic evolutionary aspects to translational applications in genetic and epigenetic diagnostics and therapy. Mutations are defined as all possible alterations in DNA sequence and sequence organization, from point mutations to genome structural variation, chromosomal aberrations and aneuploidy. Epimutations are defined as alterations in the epigenome, i.e., changes in DNA methylation, histone modification and small regulatory RNAs. MR publishes articles in the following areas: Of special interest are basic mechanisms through which DNA damage and mutations impact development and differentiation, stem cell biology and cell fate in general, including various forms of cell death and cellular senescence. The study of genome instability in human molecular epidemiology and in relation to complex phenotypes, such as human disease, is considered a growing area of importance. Mechanisms of (epi)mutation induction, for example, during DNA repair, replication or recombination; novel methods of (epi)mutation detection, with a focus on ultra-high-throughput sequencing. Landscape of somatic mutations and epimutations in cancer and aging. Role of de novo mutations in human disease and aging; mutations in population genomics. Interactions between mutations and epimutations. The role of epimutations in chromatin structure and function. Mitochondrial DNA mutations and their consequences in terms of human disease and aging. Novel ways to generate mutations and epimutations in cell lines and animal models.