Structural and Dynamical Signatures of Local DNA Damage in Live Cells

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 μm 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.