Live cell single-molecule imaging to study DNA repair in human cells.

The genetic material in human cells is continuously exposed to a wide variety of insults that can induce different DNA lesions. To maintain genomic stability and prevent potentially deleterious genetic changes caused by DNA damage, mammalian cells have evolved a number of pathways that repair specific types of DNA damage. These DNA repair pathways vary in their accuracy, some providing high-fidelity repair while others are error-prone and are only activated as a last resort. Adding additional complexity to cellular mechanisms of DNA repair is the DNA damage response which is a sophisticated a signaling network that coordinates repair outcomes, cell-cycle checkpoint activation, and cell fate decisions. As a result of the sheer complexity of the various DNA repair pathways and the DNA damage response there are large gaps in our understanding of the molecular mechanisms underlying DNA damage repair in human cells. A key unaddressed question is how the dynamic recruitment of DNA repair factors contributes to repair kinetics and repair pathway choice in human cells. Methodological advances in live cell single-molecule imaging over the last decade now allow researchers to directly observe and analyze the dynamics of DNA repair proteins in living cells with high spatiotemporal resolution. Live cell single-molecule imaging combined with single-particle tracking can provide direct insight into the biochemical reactions that control DNA repair and has the power to identify previously unobservable processes in living cells. This review summarizes the main considerations for experimental design and execution for live cell single-molecule imaging experiments and describes how they can be used to define the molecular mechanisms of DNA damage repair in mammalian cells.
Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2023 Elsevier B.V. All rights reserved.)