title: A double-strand break repair defect in ATM-deficient cells contributes to radiosensitivity.
abstract: The ATM protein, which is mutated in individuals with ataxia telangiectasia (AT), is central to cell cycle checkpoint responses initiated by DNA double-strand breaks (DSBs). ATM's role in DSB repair is currently unclear as is the basis underlying the radiosensitivity of AT cells. We applied immunofluorescence detection of gamma-H2AX nuclear foci and pulsed-field gel electrophoresis to quantify the repair of DSBs after X-ray doses between 0.02 and 80 Gy in confluence-arrested primary human fibroblasts from normal individuals and patients with mutations in ATM and DNA ligase IV, a core component of the nonhomologous end-joining (NHEJ) repair pathway. Cells with hypomorphic mutations in DNA ligase IV exhibit a substantial repair defect up to 24 h after treatment but continue to repair for several days and finally reach a level of unrepaired DSBs similar to that of wild-type cells. Additionally, the repair defect in NHEJ mutants is dose dependent. ATM-deficient cells, in contrast, repair the majority of DSBs with normal kinetics but fail to repair a subset of breaks, irrespective of the initial number of lesions induced. Significantly, after biologically relevant radiation doses and/or long repair times, the repair defect in AT cells is more pronounced than that of NHEJ mutants and correlates with radiosensitivity. NHEJ-defective cells analyzed for survival following delayed plating after irradiation show substantial recovery while AT cells fail to show any recovery. These data argue that the DSB repair defect underlies a significant component of the radiosensitivity of AT cells.
authors.affiliation: Fachrichtung Biophysik, Universität des Saarlandes, Homburg/Saar, Germany. markus.loebrich@uniklinik-saarland.de
authors.forename: Martin
authors.initials: M
authors.lastname: Kühne
authors.forename: Enriqueta
authors.initials: E
authors.lastname: Riballo
authors.forename: Nicole
authors.initials: N
authors.lastname: Rief
authors.forename: Kai
authors.initials: K
authors.lastname: Rothkamm
authors.forename: Penny A
authors.initials: PA
authors.lastname: Jeggo
authors.forename: Markus
authors.initials: M
authors.lastname: Löbrich
mesh_terms: D001260:Ataxia Telangiectasia; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins; D002460:Cell Line; D004249:DNA Damage; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000235:genetics / Q000528:radiation effects; D004260:DNA Repair / Q000235:genetics* / Q000528:radiation effects*; D004268:DNA-Binding Proteins; D004307:Dose-Response Relationship, Radiation; D016521:Electrophoresis, Gel, Pulsed-Field; D005347:Fibroblasts / Q000502:physiology / Q000528:radiation effects; D006801:Humans; D015151:Immunoblotting; D007700:Kinetics; D008168:Lung / Q000166:cytology / Q000528:radiation effects; D017346:Protein Serine-Threonine Kinases / Q000172:deficiency* / Q000235:genetics*; D011836:Radiation Tolerance; D025521:Tumor Suppressor Proteins; D014965:X-Rays
chemical_list: D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C000607893:LIG4 protein, human; D025521:Tumor Suppressor Proteins; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D011088:DNA Ligases; D000072481:DNA Ligase ATP