Double-Strand Break Clustering: An Economical and Effective Strategy for DNA Repair

TL;DR

This study reveals that non-specific DNA double-strand breaks (DSBs) cluster during early repair stages influenced by spatial proximity and phase separation, with a new dynamic model predicting clustering behavior.

Contribution

It introduces a high-precision microscopy method to study DSB clustering, and develops a Monte Carlo model to predict the spatiotemporal dynamics of DSB clustering during DNA repair.

Findings

DSB clustering occurs during early DDR stages.

Clustering probability is constant at 0.8-1.4 um distances.

A dynamic model accurately predicts clustering behavior.

Abstract

In mammalian cells, repair centers for DNA double-strand breaks (DSBs) have been identified. However, previous researches predominantly rely on methods that induce specific DSBs by cutting particular DNA sequences. The clustering and its spatiotemporal properties of non-specifically DSBs, especially those induced by environmental stresses such as irradiation, remains unclear. In this study, we used Dragonfly microscopy to induce high-precision damage in cells and discovered that DSB clustering during the early stages of DNA damage response (DDR) and repair, but not during the repair plateau phase. Early in DDR, DSB clustered into existing 53BP1 foci. The DSB clustering at different stages has different implications for DNA repair. By controlling the distance between adjacent damage points, we found that the probability of DSB clustering remains constant at distances of 0.8 - 1.4 um, while clustering does not occur beyond 1.4 um. Within the 0.8 um range, the probability of clustering significantly increases due to the phase separation effect of 53BP1. Using a Monte Carlo approach, we developed a dynamic model of 53BP1 foci formation, fission, and fusion. This model accurately predicts experimental outcomes and further demonstrates the temporal and spatial influences on DSB clustering. These results showed that, similarly to specifically induced DSBs, non-specifically induced DSBs can also cluster. The extent of DSB clustering is influenced by both temporal and spatial factors, which provide new insights into the dynamics of DSB clustering and the role of 53BP1 in DNA repair processes. Such findings could enhance our understanding of DNA damage responses and help us improve DNA repair therapies in disease.