Agarwal Harsha, Reisser Matthias, Wortmann Celina, Gebhardt J Christof M
Institute of Biophysics, Ulm University, Ulm, Germany.
Institute of Biophysics, Ulm University, Ulm, Germany.
Biophys J. 2017 May 23;112(10):2051-2055. doi: 10.1016/j.bpj.2017.04.018. Epub 2017 May 6.
The three-dimensional arrangement of chromatin encodes regulatory traits important for nuclear processes such as transcription and replication. Chromatin topology is in part mediated by the architectural protein CCCTC-binding factor (CTCF) that binds to the boundaries of topologically associating domains. Whereas sites of CTCF interactions are well characterized, little is known on how long CTCF binds to chromatin and how binding evolves during the cell cycle. We monitored CTCF-chromatin interactions by live cell single molecule tracking in different phases of the cell cycle. In G1-, S-, and G2-phases, a majority of CTCF molecules was bound transiently (∼0.2 s) to chromatin, whereas minor fractions were bound dynamically (∼4 s) or stably (>15 min). During mitosis, CTCF was mostly excluded from chromatin. Our data suggest that CTCF scans DNA in search for two different subsets of specific target sites and provide information on the timescales over which topologically associating domains might be restructured. During S-phase, dynamic and stable interactions decreased considerably compared to G1-phase, but were resumed in G2-phase, indicating that specific interactions need to be dissolved for replication to proceed.
染色质的三维排列编码了对转录和复制等核过程至关重要的调控特征。染色质拓扑结构部分由与拓扑相关结构域边界结合的结构蛋白CCCTC结合因子（CTCF）介导。虽然CTCF相互作用位点已得到充分表征，但对于CTCF与染色质结合多长时间以及结合在细胞周期中如何演变知之甚少。我们通过活细胞单分子追踪监测了细胞周期不同阶段的CTCF-染色质相互作用。在G1期、S期和G2期，大多数CTCF分子与染色质短暂结合（约0.2秒），而少数部分动态结合（约4秒）或稳定结合（>15分钟）。在有丝分裂期间，CTCF大多被排除在染色质之外。我们的数据表明，CTCF扫描DNA以寻找特定靶位点的两个不同子集，并提供有关拓扑相关结构域可能重组的时间尺度的信息。在S期，与G1期相比，动态和稳定相互作用显著减少，但在G2期恢复，这表明特定相互作用需要解除才能进行复制。
