Structural and Dynamical Signatures of Local DNA Damage in Live Cells
Structural and Dynamical Signatures of Local DNA Damage in Live Cells
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DOI：
10.1016/j.bpj.2019.10.042
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发表时间：
2020-05-05
影响因子：
3.4
通讯作者：
Zidovska, Alexandra
中科院分区：
文献类型：
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作者：
Eaton, Jonah A.;Zidovska, Alexandra
细胞核内染色质的动态组织在基因调控和基因组复制以及维持基因组完整性中起着关键作用。尽管基因组的静态折叠状态已被广泛研究，但转录或DNA修复等过程的动态特征仍然是一个悬而未决的问题。在这里，我们研究了人类细胞中响应局部DNA损伤的间期染色质动力学，特别是DNA双链断裂（DSBs）。同时使用双色旋转圆盘共聚焦显微镜，我们监测DSB动态和周围染色质的压实，分别通过荧光标记53BP1和组蛋白H2B进行可视化。我们的研究揭示了位于核内部和外围（距离核膜小于1 μ m）的dsb的移动性之间的惊人差异，其中内部dsb的移动性几乎是外围dsb的两倍。值得注意的是，我们发现DSB位点以独特的染色质压实剖面的形式具有强大的结构特征。此外，我们的数据表明，DSB运动是亚弥漫性的和atp依赖的，并表现出独特的动态特征，不同于未受损的染色质。我们的研究结果表明，DSB迁移率遵循一种普遍的关系，这种关系仅由描述DSB及其局部环境的物理参数定义，例如DSB焦点大小（由53BP1的局部积累表示），DSB密度和局部染色质压实。这表明DSB相关的修复过程是稳健的，并且可能是确定的，因为观察到的动态特征（DSB迁移率）可以仅仅通过它们的结构特征（DSB焦点大小，局部染色质压实）来解释。这些知识可能有助于检测活细胞中的局部DNA损伤，并有助于我们对健康和疾病中基因组完整性的生物物理学理解。
The dynamic organization of chromatin inside the cell nucleus plays a key role in gene regulation and genome replication, as well as maintaining genome integrity. Although the static folded state of the genome has been extensively studied, dynamical signatures of processes such as transcription or DNA repair remain an open question. Here, we investigate the interphase chromatin dynamics in human cells in response to local DNA damage, specifically, DNA double-strand breaks (DSBs). Using simultaneous two-color spinning-disk confocal microscopy, we monitor the DSB dynamics and the compaction of the surrounding chromatin, visualized by fluorescently labeled 53BP1 and histone H2B, respectively. Our study reveals a surprising difference between the mobility of DSBs located in the nuclear interior versus periphery (less than 1 mu m from the nuclear envelope), with the interior DSBs being almost twice as mobile as the periphery DSBs. Remarkably, we find that the DSB sites possess a robust structural signature in a form of a unique chromatin compaction profile. Moreover, our data show that the DSB motion is subdiffusive and ATP-dependent and exhibits unique dynamical signatures, different from those of undamaged chromatin. Our findings reveal that the DSB mobility follows a universal relationship defined solely by the physical parameters describing the DSBs and their local environment, such as the DSB focus size (represented by the local accumulation of 53BP1), DSB density, and the local chromatin compaction. This suggests that the DSB-related repair processes are robust and likely deterministic because the observed dynamical signatures (DSB mobility) can be explained solely by their structural features (DSB focus size, local chromatin compaction). Such knowledge might help in detecting local DNA damage in live cells, as well as in aiding our biophysical understanding of genome integrity in health and disease.