Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology, Higashi 1-1-1, Tsukuba 305-8566, Japan.
Nucleic Acids Res. 2012 Oct;40(19):9482-92. doi: 10.1093/nar/gks779. Epub 2012 Aug 16.
The Bmal1 gene is essential for the circadian system, and its promoter has a unique open chromatin structure. We examined the mechanism of topoisomerase I (Top1) to understand the role of the unique chromatin structure in Bmal1 gene regulation. Camptothecin, a Top1 inhibitor, and Top1 small interfering RNA (siRNA) enhanced Baml1 transcription and lengthened its circadian period. Top1 is located at an intermediate region between two ROREs that are critical cis-elements of circadian transcription and the profile of Top1 binding indicated anti-phase circadian oscillation of Bmal1 transcription. Promoter assays showed that the Top1-binding site is required for transcriptional suppression and that it functions cooperatively with the distal RORE, supporting that Bmal1 transcription is upregulated by Top1 inhibition. A DNA fragment between the ROREs, where the Top1-binding site is located, behaved like a right-handed superhelical twist, and modulation of Top1 activity by camptothecin and Top1 siRNA altered the footprint profile, indicating modulation of the chromatin structure. These data indicate that Top1 modulates the chromatin structure of the Bmal1 promoter, regulates Bmal1 transcription and influences the circadian period.
Bmal1 基因是生物钟系统的必需基因，其启动子具有独特的开放染色质结构。我们研究了拓扑异构酶 I（Top1）的作用机制，以了解这种独特的染色质结构在 Bmal1 基因调控中的作用。拓扑异构酶 I 抑制剂喜树碱和 Top1 小干扰 RNA（siRNA）增强了 Baml1 的转录，并延长了其生物钟周期。Top1 位于两个 RORE 之间的一个中间区域，RORE 是生物钟转录的关键顺式元件，Top1 结合的特征表明 Bmal1 转录呈反相生物钟振荡。启动子分析表明，Top1 结合位点对于转录抑制是必需的，并且它与远端 RORE 协同作用，支持 Top1 抑制可上调 Bmal1 转录。位于 Top1 结合位点的 RORE 之间的 DNA 片段表现为右手超螺旋扭曲，喜树碱和 Top1 siRNA 对 Top1 活性的调节改变了足迹特征，表明染色质结构的调节。这些数据表明，Top1 调节 Bmal1 启动子的染色质结构，调节 Bmal1 转录并影响生物钟周期。
