title: Distinct roles of XRCC4 and Ku80 in non-homologous end-joining of endonuclease- and ionizing radiation-induced DNA double-strand breaks.
abstract: Non-homologous end-joining (NHEJ) of DNA double-strand breaks (DSBs) is mediated by two protein complexes comprising Ku80/Ku70/DNA-PKcs/Artemis and XRCC4/LigaseIV/XLF. Loss of Ku or XRCC4/LigaseIV function compromises the rejoining of radiation-induced DSBs and leads to defective V(D)J recombination. In this study, we sought to define how XRCC4 and Ku80 affect NHEJ of site-directed chromosomal DSBs in murine fibroblasts. We employed a recently developed reporter system based on the rejoining of I-SceI endonuclease-induced DSBs. We found that the frequency of NHEJ was reduced by more than 20-fold in XRCC4-/- compared to XRCC4+/+ cells, while a Ku80 knock-out reduced the rejoining efficiency by only 1.4-fold. In contrast, lack of either XRCC4 or Ku80 increased end degradation and shifted repair towards a mode that used longer terminal microhomologies for rejoining. However, both proteins proved to be essential for the repair of radiation-induced DSBs. The remarkably different phenotype of XRCC4- and Ku80-deficient cells with regard to the repair of enzyme-induced DSBs mirrors the embryonic lethality of XRCC4 knock-out mice as opposed to the viability of the Ku80 knock-out. Thus, I-SceI-induced breaks may resemble DSBs arising during normal DNA metabolism and mouse development. The removal of these breaks likely has different genetic requirements than the repair of radiation-induced DSBs.
authors.affiliation: Laboratory of Radiobiology & Experimental Radiation Oncology, Department of Radiotherapy and Radiation Oncology, University Medical School Hamburg-Eppendorf, Martinistrasse 52, D-20246 Hamburg, Germany.
authors.forename: Leonie
authors.initials: L
authors.lastname: Schulte-Uentrop
authors.forename: Raafat A
authors.initials: RA
authors.lastname: El-Awady
authors.forename: Lena
authors.initials: L
authors.lastname: Schliecker
authors.forename: Henning
authors.initials: H
authors.lastname: Willers
authors.forename: Jochen
authors.initials: J
authors.lastname: Dahm-Daphi
mesh_terms: D000818:Animals; D034961:Antigens, Nuclear / Q000235:genetics / Q000502:physiology*; D002478:Cells, Cultured; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D017353:Gene Deletion; D000072200:Ku Autoantigen; D051379:Mice; D011836:Radiation Tolerance; D011839:Radiation, Ionizing; D029701:Saccharomyces cerevisiae Proteins
chemical_list: D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; C511788:XRCC4 protein, mouse; C037725:SCEI protein, S cerevisiae; D015252:Deoxyribonucleases, Type II Site-Specific; C000606510:Xrcc6 protein, mouse; D000072200:Ku Autoantigen