Topoisomerase II-induced Chromosome Breakage and Translocation Is Determined by Chromosome Architecture and Transcriptional Activity [Nascent RNA]

Topoisomerase II (TOP2) relieves torsional stress during transcription, DNA replication and chromosome segregation, by forming transient cleavage complex intermediates (TOP2ccs) that contain TOP2-linked DNA breaks. While TOP2ccs are normally reversible they can be ‘trapped’ by chemotherapeutic drugs such as etoposide, and subsequently converted into irreversible TOP2-linked DSBs that threaten genome stability. Here, using genomics approaches, we have quantified the etoposide-induced trapping of TOP2ccs, their conversion into irreversible TOP2-linked DSBs, and their processing during DNA repair genome-wide, as a function of time. We find that while TOP2 trapping is independent of transcription it requires pre-existing binding of cohesin to DNA. In contrast, the conversion of trapped TOP2ccs to irreversible DSBs during DNA repair is accelerated two-fold at transcribed loci, relative to non-transcribed loci. This conversion is dependent on proteasomal degradation and TDP2 phosphodiesterase activity. Quantitative modeling shows that only two critical features of pre-existing chromatin structure- namely, cohesin binding and transcriptional activity- can be used to accurately predict the kinetics of TOP2-induced DSBs. Thus, our study permits a mechanistic understanding of TOP2 induced genome instability.

拓扑异构酶II（Topoisomerase II, TOP2）可通过组装包含TOP2连接型DNA断裂的瞬时切割复合物中间体（TOP2ccs），在转录、DNA复制与染色体分离过程中缓解DNA扭转应力。正常状态下TOP2cc具备可逆性，但可被依托泊苷等化疗药物“捕获”，进而转化为威胁基因组稳定性的不可逆TOP2连接型DNA双链断裂（DNA double-strand breaks, DSBs）。本研究借助基因组学技术手段，定量分析了依托泊苷诱导的TOP2cc捕获事件、其向不可逆TOP2连接型DSBs的转化过程，以及全基因组范围内DNA修复进程中此类复合物的动态加工过程，并结合时间维度系统考察了相关动态变化。研究发现，TOP2的捕获过程不依赖转录活性，但需要黏连蛋白（cohesin）预先结合至DNA分子。与之形成对比的是，相较于非转录位点，转录位点处捕获的TOP2cc向不可逆DSBs的转化速率提升了两倍。该转化过程依赖蛋白酶体降解（proteasomal degradation）途径与TDP2磷酸二酯酶（TDP2 phosphodiesterase）的活性。定量建模结果表明，仅需预先存在的染色质结构的两项关键特征——黏连蛋白结合与转录活性——即可精准预测TOP2诱导DSBs的动力学过程。综上，本研究实现了对TOP2诱导基因组不稳定机制的系统性阐释。