Topoisomerases I and II facilitate the translocation of condensin DC in C. elegans [ChIP-Seq II]. Topoisomerases I and II facilitate the translocation of condensin DC in C. elegans [ChIP-Seq II]

Condensin complexes are evolutionarily conserved molecular motors from the structural maintenance of chromosomes (SMC) family, that use ATPase activity to translocate along DNA and form loops. Condensin and topoisomerase II (TOP-2) are essential for the structure and function of mitotic chromosomes. While condensin-mediated DNA looping is thought to direct TOP-2 chain-passing activity to separate sister chromatids, it is not known if TOP-2 in turn regulates loop formation. Here we used an X chromosome specific condensin that represses transcription for dosage compensation in C.elegans, to determine how DNA topology affects SMC translocation in vivo. We applied auxin-inducible degradation of topoisomerases I and II to determine their effect on condensin DC binding and function. We found that both topoisomerases colocalize with condensin DC and control its movement at different genomic scales. TOP-2 depletion hindered condensin DC spreading over long distances, resulting in accumulation around its X-specific recruitment sites and shorter Hi-C interactions. In contrast, TOP-1 depletion did not affect long-range spreading but resulted in accumulation of condensin DC within gene bodies, specially of highly expressed and long genes. Both TOP-1 and TOP-2 depletion resulted in X chromosome upregulation indicating that condensin DC translocation at both scales is required for its function in transcriptional repression. Together our work reveals distinct DNA topological requirements for two modes of condensin DC association with chromatin: long-range linear translocation that requires decatenation and unknotting of DNA and short-range binding to genes that requires resolution of transcription-induced supercoiling. Overall design: ChIP-seq data in WT, controls and topoisomerases I and II depleted conditions . Depletion was performed by Auxin inducible degradation. All experiments were performed in two biological replicates. L2/L3 larvae were used for most experiments. TOP-2 ChIP-seq in embryos and adults are also included.

凝缩蛋白复合物（Condensin complex）是进化保守的结构维持染色体（structural maintenance of chromosomes, SMC）家族分子马达，通过ATP酶活性沿DNA移位并形成DNA环。凝缩蛋白与拓扑异构酶II（topoisomerase II, TOP-2）对有丝分裂染色体的结构与功能至关重要。以往研究认为，凝缩蛋白介导的DNA环化可指导TOP-2的链传递活性以分离姐妹染色单体，但目前尚不清楚TOP-2是否可反向调控环的形成。
本研究利用秀丽隐杆线虫（C. elegans）中参与剂量补偿、抑制转录的X染色体特异性凝缩蛋白DC（condensin DC），探究体内DNA拓扑结构对SMC移位的调控作用。我们通过生长素诱导降解（auxin-inducible degradation）系统靶向降解拓扑异构酶I与II，以明确二者对凝缩蛋白DC的结合与功能的影响。
研究发现，两种拓扑异构酶均与凝缩蛋白DC共定位，并在不同基因组尺度上调控其运动。TOP-2耗竭会阻碍凝缩蛋白DC的长距离扩散，导致其在X染色体特异性招募位点周围积累，并缩短Hi-C互作的长度范围。与之相反，TOP-1耗竭不会影响长距离扩散，但会使凝缩蛋白DC在基因本体（gene bodies）内发生积累，尤其富集于高表达的长基因中。TOP-1与TOP-2的耗竭均会导致X染色体转录上调，表明两种尺度下的凝缩蛋白DC移位均是其实现转录抑制功能的必要条件。
综上，本研究揭示了凝缩蛋白DC与染色质结合的两种模式对DNA拓扑结构的差异化需求：一是依赖DNA解连环与消结的长距离线性移位模式，二是需要解决转录诱导超螺旋的基因短程结合模式。
整体实验设计：野生型（WT）、对照组及拓扑异构酶I、II耗竭条件下的ChIP-seq测序数据。耗竭操作通过生长素诱导降解系统完成。所有实验均设置2个生物学重复。多数实验采用L2/L3期幼虫作为实验材料，同时还包含胚胎与成虫阶段的TOP-2 ChIP-seq数据。