title: Clustered DNA damage induces pan-nuclear H2AX phosphorylation mediated by ATM and DNA-PK.
abstract: DNA double-strand breaks (DSB) are considered as the most deleterious DNA lesions, and their repair is further complicated by increasing damage complexity. However, the molecular effects of clustered lesions are yet not fully understood. As the locally restricted phosphorylation of H2AX to form γH2AX is a key step in facilitating efficient DSB repair, we investigated this process after localized induction of clustered damage by ionizing radiation. We show that in addition to foci at damaged sites, H2AX is also phosphorylated in undamaged chromatin over the whole-cell nucleus in human and rodent cells, but this is not related to apoptosis. This pan-nuclear γH2AX is mediated by the kinases ataxia telangiectasia mutated and DNA-dependent protein kinase (DNA-PK) that also phosphorylate H2AX at DSBs. The pan-nuclear response is dependent on the amount of DNA damage and is transient even under conditions of impaired DSB repair. Using fluorescence recovery after photobleaching (FRAP), we found that MDC1, but not 53BP1, binds to the nuclear-wide γH2AX. Consequently, the accumulation of MDC1 at DSBs is reduced. Altogether, we show that a transient dose-dependent activation of the kinases occurring on complex DNA lesions leads to their nuclear-wide distribution and H2AX phosphorylation, yet without eliciting a full pan-nuclear DNA damage response.
authors.affiliation: Department of Biophysics, GSI Helmholtz Center for Heavy Ion Research, Planckstrasse 1, D-64291 Darmstadt, Germany.
authors.forename: Barbara
authors.initials: B
authors.lastname: Meyer
authors.forename: Kay-Obbe
authors.initials: KO
authors.lastname: Voss
authors.forename: Frank
authors.initials: F
authors.lastname: Tobias
authors.forename: Burkhard
authors.initials: B
authors.lastname: Jakob
authors.forename: Marco
authors.initials: M
authors.lastname: Durante
authors.forename: Gisela
authors.initials: G
authors.lastname: Taucher-Scholz
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002460:Cell Line; D002467:Cell Nucleus / Q000201:enzymology* / Q000378:metabolism / Q000528:radiation effects; D002478:Cells, Cultured; D002843:Chromatin / Q000737:chemistry; D006224:Cricetinae; D053903:DNA Breaks, Double-Stranded; D051747:DNA-Activated Protein Kinase / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006657:Histones / Q000032:analysis / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011839:Radiation, Ionizing; D015534:Trans-Activators / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000378:metabolism*; D000071857:Tumor Suppressor p53-Binding Protein 1
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D006657:Histones; D047908:Intracellular Signaling Peptides and Proteins; C413163:MDC1 protein, human; D009687:Nuclear Proteins; C498301:TP53BP1 protein, human; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins; D000071857:Tumor Suppressor p53-Binding Protein 1; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C576325:Atm protein, mouse; D051747:DNA-Activated Protein Kinase; D017346:Protein Serine-Threonine Kinases