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Topoisomerase IIalpha-dependent induction of a persistent DNA damage response in response to transient etoposide

Sébastien Soubeyrand1, Louise Pope, Robert J G Haché

  • 1The Ottawa Health Research Institute, University of Ottawa, 451 Smyth Road, Ottawa, ON K1H 8M5, Canada.

Molecular Oncology
|October 28, 2009
PubMed

Insights

Etoposide causes rapid, persistent DNA damage by trapping topoisomerase II (topoII) in DNA, leading to cell death even before DNA replication. This damage occurs in open chromatin and requires functional topoII to resolve.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Genetics

Background:

  • Etoposide, a topoisomerase II (topoII) poison, is known to cause cytotoxicity by trapping topoII in DNA, leading to lethal lesions during DNA replication.
  • However, rapid cell death following short-term etoposide treatment suggests that cytotoxic lesions form before DNA replication begins.

Purpose of the Study:

  • To investigate the rapid DNA damage response and cytotoxicity induced by etoposide, independent of DNA replication.
  • To elucidate the role of topoisomerase IIalpha (topoIIalpha) in etoposide-induced DNA damage and cell death.

Main Methods:

  • Treatment of cells with etoposide and subsequent analysis of DNA damage response markers (ATM, MDC1, DNA-PKcs, gammaH2AX).
  • Assessment of DNA repair dynamics upon etoposide withdrawal.
  • Utilizing siRNA to ablate topoisomerase II (topoII) expression.
  • Investigating the effect of DNA replication/transcription inhibition and histone deacetylase inhibition (trichostatin A) on gammaH2AX induction.

Main Results:

  • Short-term etoposide treatment induced a rapid and significant double-stranded DNA damage response, marked by gammaH2AX foci, which persisted despite DNA repair.
  • TopoII siRNA ablation demonstrated that etoposide toxicity and gammaH2AX response are linked to impaired resolution of topoIIalpha-mediated DNA damage.
  • GammaH2AX induction was independent of DNA replication and transcription but was enhanced by trichostatin A, suggesting a role for chromatin structure.

Conclusions:

  • Etoposide lethality is associated with the rapid generation of persistent DNA defects dependent on topoIIalpha.
  • These defects occur within topologically open chromatin domains and are processed by the DNA damage response machinery.
  • The findings highlight a critical role for topoIIalpha in resolving etoposide-induced DNA damage, independent of the cell cycle stage.

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