Early effects of topoisomerase I inhibition on RNA polymerase II along transcribed genes in human cells
Early effects of topoisomerase I inhibition on RNA polymerase II along transcribed genes in human cells
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DOI：
10.1016/j.jmb.2005.12.069
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发表时间：
2006-03-17
影响因子：
5.6
通讯作者：
Capranico, G
中科院分区：
文献类型：
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作者：
Khobta, A;Ferri, F;Capranico, G
我们已经通过染色质免疫沉淀（ChIP）确定了喜树碱和a-amanitin对RNA聚合酶II （RNAPII）、tata结合蛋白（TBP）、DNA拓扑异构酶I （Top1）和人类转录位点组蛋白成分的基因组DNA结合位点的早期影响。这两种药物引起活性染色质明显不同的改变。喜树碱诱导启动子暂停位点的RNAPII密度特异性降低，组蛋白修饰表明染色质可及性增加。a- amanitin导致RNAPII在转录基因上的积累，与染色质结合的TBP减少，染色质结构不易接近。有趣的是，启动子暂停位点的RNAPII减少发生在喜树碱治疗后的5-10分钟内，并且不是对复制依赖性DNA断裂的反应。RNAPII沿转录基因的ChIP分析表明，RNAPII水平在内部外显子处短暂升高，并且喜树碱的作用可以被cdk抑制剂DRB完全逆转。Top1被发现在活性染色质中富集，因此表明Top1在转录模板和/或邻近调节区域的抑制会立即影响活性基因上的RNAPII。这些发现是喜树碱对RNAPII结合到转录基因组区域作用的新的体内证据，并且与Top1活性可以在启动子清除水平上参与转录调节的假设一致。(c) 2005 Elsevier Ltd版权所有。
We have determined the early effects of camptothecin and a-amanitin on genomic DNA-binding sites of RNA polymerase II (RNAPII), TATA-binding protein (TBP), DNA topoisomerase I (Top1), and histone components in human transcribed loci by chromatin-immunoprecipitation (ChIP). The two agents caused notably different alterations in active chromatin. Camptothecin induced a specific reduction of RNAPII density at promoter pause sites and histone modifications suggesting an increased chromatin accessibility. a-Amanitin caused an accumulation of RNAPII at transcribed genes, a reduction of TBP bound to chromatin and a less accessible chromatin structure. Interestingly, RNAPII reduction at promoter pause sites occurred within 5-10 min of camptothecin treatment, and was not a response to replication-dependent DNA breaks. ChIP analyses of RNAPII along transcribed genes indicated that RNAPII levels were transiently increased at internal exons, and that camptothecin effects could be fully reversed by DRB, a cdk inhibitor. Top1 was found to be enriched in active chromatin, therefore suggesting that Top1 inhibition at the transcribed template and/or adjacent regulating regions immediately affects RNAPII at active genes. The findings are novel in vivo evidence of camptothecin effects on RNAPII bound to transcribing genomic regions, and are consistent with the hypothesis that Top1 activity can be involved in transcription regulation at the level of promoter clearance. (c) 2005 Elsevier Ltd. All rights reserved.