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A protease pathway for the repair of topoisomerase II-DNA covalent complexes

Ailing Zhang1, Yi Lisa Lyu, Chao-Po Lin

  • 1Department of Pharmacology, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.

The Journal of Biological Chemistry
|September 16, 2006
PubMed

Insights

The proteasome aids in repairing DNA double-strand breaks (DSBs) by degrading topoisomerase II (TopII)-DNA adducts. Inhibiting the proteasome reduces VP-16-induced DSBs, highlighting its role in processing these specific DNA lesions.

Area of Science:

  • Molecular Biology
  • DNA Repair Mechanisms
  • Cellular Proteostasis

Background:

  • Protein-DNA adducts, particularly topoisomerase II (TopII)-DNA complexes, are challenging DNA lesions.
  • Previous research suggests the proteasome degrades TopII-DNA adducts, potentially exposing DNA double-strand breaks (DSBs).
  • The precise role of the proteasome in processing TopII-DNA adducts and subsequent DNA repair remains incompletely understood.

Purpose of the Study:

  • To investigate the involvement of the proteasome in processing topoisomerase II (TopII)-DNA covalent complexes into DNA double-strand breaks (DSBs).
  • To determine if proteasome inhibition affects the formation of VP-16 (etoposide)-induced DSBs and DNA damage signaling.
  • To elucidate the specificity of the proteasome pathway in the repair of TopII-DNA adducts versus other DNA damage types.

Main Methods:

  • Induction of TopII-DNA covalent complexes using the anticancer drug VP-16 (etoposide) in mammalian cells.
  • Treatment with the proteasome inhibitor MG132 to assess the impact on DNA damage.
  • Utilized top2beta knock-out mouse embryonic fibroblasts, Top2beta small interfering RNA knockdown PC12 cells, and postmitotic neurons (lacking TopIIalpha).
  • Monitored VP-16-induced DSBs using neutral comet assay and DNA damage signaling via gamma-H2AX.
  • Compared the effect of proteasome inhibition on VP-16-induced damage versus hydrogen peroxide-induced DNA damage.

Main Results:

  • VP-16-induced DNA double-strand breaks (DSBs) and gamma-H2AX signaling were significantly reduced by the proteasome inhibitor MG132.
  • Proteasome inhibition attenuated VP-16-induced DNA damage signals in cells lacking TopIIbeta or TopIIalpha, indicating involvement in both TopII isoforms.
  • Hydrogen peroxide-induced gamma-H2AX levels were unaffected by proteasome inhibition, demonstrating pathway specificity.
  • These findings support the hypothesis that proteasomal degradation processes TopII-DNA adducts, facilitating subsequent DNA repair.

Conclusions:

  • The proteasome plays a crucial role in processing topoisomerase II (TopII)-DNA covalent complexes into DNA double-strand breaks (DSBs).
  • This proteasome-dependent processing is essential for the efficient repair of TopII-DNA adducts induced by agents like VP-16.
  • The proteasome specifically functions in the repair pathway for TopII-DNA adducts, distinct from general oxidative DNA damage response.

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