The dynamics of Ku70/80 and DNA-PKcs at DSBs induced by ionizing radiation is dependent on the complexity of damage

Nucleic Acids Res. 2012 Nov;40(21):10821-31. doi: 10.1093/nar/gks879. Epub 2012 Sep 24.

Abstract

DNA double-strand breaks (DSBs) are biologically one of the most important cellular lesions and possess varying degrees of chemical complexity. The notion that the repairability of more chemically complex DSBs is inefficient led to the concept that the extent of DSB complexity underlies the severity of the biological consequences. The repair of DSBs by non-homologous end joining (NHEJ) has been extensively studied but it remains unknown whether more complex DSBs require a different sub-set of NHEJ protein for their repair compared with simple DSBs. To address this, we have induced DSBs in fluorescently tagged mammalian cells (Ku80-EGFP, DNA-PKcs-YFP or XRCC4-GFP, key proteins in NHEJ) using ultra-soft X-rays (USX) or multi-photon near infrared (NIR) laser irradiation. We have shown in real-time that simple DSBs, induced by USX or NIR microbeam irradiation, are repaired rapidly involving Ku70/80 and XRCC4/Ligase IV/XLF. In contrast, DSBs with greater chemical complexity are repaired slowly involving not only Ku70/80 and XRCC4/Ligase IV/XLF but also DNA-PKcs. Ataxia telangiectasia-mutated inhibition only retards repair of the more chemically complex DSBs which require DNA-PKcs. In summary, the repair of DSBs by NHEJ is highly regulated with pathway choice and kinetics of repair dependent on the chemical complexity of the DSB.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Antigens, Nuclear / metabolism*
  • Ataxia Telangiectasia Mutated Proteins
  • Cell Cycle Proteins / antagonists & inhibitors
  • Cell Line
  • DNA Breaks, Double-Stranded*
  • DNA End-Joining Repair*
  • DNA-Activated Protein Kinase / metabolism*
  • DNA-Binding Proteins / antagonists & inhibitors
  • DNA-Binding Proteins / metabolism*
  • Kinetics
  • Ku Autoantigen
  • Protein Serine-Threonine Kinases / antagonists & inhibitors
  • Radiation, Ionizing*
  • Tumor Suppressor Proteins / antagonists & inhibitors

Substances

  • Antigens, Nuclear
  • Cell Cycle Proteins
  • DNA-Binding Proteins
  • Tumor Suppressor Proteins
  • Ataxia Telangiectasia Mutated Proteins
  • DNA-Activated Protein Kinase
  • Protein Serine-Threonine Kinases
  • Ku Autoantigen