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Publications

Journal Article Research Support, N.I.H., Intramural Research Support, N.I.H., Extramural Research Support, U.S. Gov't, Non-P.H.S. Ppublish 5 citations

PubMed: 37670571 DOI: 10.1093/nar/gkad735

Real-time imaging of drug-induced trapping of cellular topoisomerases and poly(ADP-ribose) polymerase 1 at the single-molecule level.

Nucleic Acids Res 51(18):e97

Authors: Yilun Sun, Jiji Chen, Yves Pommier

Affiliations

  • Laboratory of Molecular Pharmacology and Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
  • Advanced Imaging and Microscopy (AIM) Resource, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892, USA.

Abstract

Topoisomerases (TOP1, TOP2α, and β) are nuclear enzymes crucial for virtually all aspects of DNA metabolisms. They also are the targets of important anti-tumor chemotherapeutics that act by trapping the otherwise reversible topoisomerase-DNA covalent complex intermediates (TOPccs) that are formed during their catalytic reactions, resulting in long-lived topoisomerase DNA-protein crosslinks (TOP-DPCs) that interfere with DNA transactions. The Poly(ADP-ribose) polymerase (PARP) family protein PARP1 is activated by DNA damage to recruit DNA repair proteins, and PARP inhibitors are another class of commonly used chemotherapeutics, which bind and trap PARP molecules on DNA. To date, the trapping of TOPccs and PARP by their respective inhibitors can only be measured by immune-biochemical methods in cells. Here, we developed an imaging-based approach enabling real-time monitoring of drug-induced trapping of TOPccs and PARP1 in live cells at the single-molecule level. Capitalizing on this approach, we calculated the fraction of self-fluorescence tag-labeled topoisomerases and PARP single-molecules that are trapped by their respective inhibitors in real time. This novel technique should help elucidate the molecular processes that repair TOPcc and PARP trapping and facilitate the development of novel topoisomerase and PARP inhibitor-based therapies.

MeSH

Dna, Pharmaceutical Preparations, Nucleic Acids, Cells, Poly Adenosine Diphosphate Ribose, Poly(Adp-Ribose) Polymerases, Poly (Adp-Ribose) Polymerase-1, Poly(Adp-Ribose) Polymerase Inhibitors

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