title: Most hydrogen peroxide-induced histone H2AX phosphorylation is mediated by ATR and is not dependent on DNA double-strand breaks.
abstract: The nuclear foci of phosphorylated histone H2AX (γH2AX) are frequently used as a marker for DNA double-strand breaks (DSBs) following ionizing radiation (IR). However, recent studies reported that γH2AX foci do not necessarily correlate with DSBs under other conditions. We showed that γH2AX foci induced by oxidative stress in hydrogen peroxide (H2O2)-treated cells displayed several different features from those induced by IR. The magnitude of γH2AX induction was heterogeneous among H2O2-treated cells. Some cells expressed small discrete γH2AX foci, whereas others expressed a gross γH2AX signal that was distributed throughout the nucleus. Oxidative stress-induced γH2AX was eliminated in DSB repair-deficient mutant cells as efficiently as in wild-type cells and was not necessarily accompanied by phosphorylated ataxia telangiectasia mutated (ATM) or 53BP1 foci. Analyses using specific inhibitors showed that ATM- and Rad3-related (ATR), rather than ATM, was the prominent kinase mediating the oxidative stress response. These results suggest that a major fraction of γH2AX induced by oxidative stress is not associated with DSBs. Single-stranded DNA arisen from stalled replication forks can cause the ATR-mediated induction of γH2AX. However, oxidative stress appeared to induce γH2AX in both S- and non-S-phase cells. These results suggest that there may be another pathway leading to the ATR-mediated induction of γH2AX in non-S-phase cells without DSBs.
authors.affiliation: Reseach Center for Radiation Protection, Fukushima Project Headquarters, Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba 263-8555, Japan and Department of Radiation Hazard Evaluation, Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300192, China tkatsu@nirs.go.jp.
authors.forename: Takanori
authors.initials: T
authors.lastname: Katsube
authors.affiliation: Reseach Center for Radiation Protection, Fukushima Project Headquarters, Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba 263-8555, Japan and Department of Radiation Hazard Evaluation, Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300192, China.
authors.forename: Masahiko
authors.initials: M
authors.lastname: Mori
authors.affiliation: Reseach Center for Radiation Protection, Fukushima Project Headquarters, Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba 263-8555, Japan and Department of Radiation Hazard Evaluation, Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300192, China.
authors.forename: Hideo
authors.initials: H
authors.lastname: Tsuji
authors.affiliation: Reseach Center for Radiation Protection, Fukushima Project Headquarters, Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba 263-8555, Japan and Department of Radiation Hazard Evaluation, Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300192, China.
authors.forename: Tadahiro
authors.initials: T
authors.lastname: Shiomi
authors.affiliation: Reseach Center for Radiation Protection, Fukushima Project Headquarters, Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba 263-8555, Japan and Department of Radiation Hazard Evaluation, Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300192, China.
authors.forename: Bing
authors.initials: B
authors.lastname: Wang
authors.affiliation: Reseach Center for Radiation Protection, Fukushima Project Headquarters, Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba 263-8555, Japan and Department of Radiation Hazard Evaluation, Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300192, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Liu
authors.affiliation: Reseach Center for Radiation Protection, Fukushima Project Headquarters, Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba 263-8555, Japan and Department of Radiation Hazard Evaluation, Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300192, China.
authors.forename: Mitsuru
authors.initials: M
authors.lastname: Nenoi
authors.affiliation: Reseach Center for Radiation Protection, Fukushima Project Headquarters, Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba 263-8555, Japan and Department of Radiation Hazard Evaluation, Institute of Radiation Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin 300192, China.
authors.forename: Makoto
authors.initials: M
authors.lastname: Onoda
mesh_terms: D064007:Ataxia Telangiectasia Mutated Proteins / Q000378:metabolism / Q000502:physiology; D002467:Cell Nucleus / Q000378:metabolism; D002470:Cell Survival; D053903:DNA Breaks, Double-Stranded; D004261:DNA Replication; D045325:HCT116 Cells; D006657:Histones / Q000378:metabolism*; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D018384:Oxidative Stress; D011499:Protein Processing, Post-Translational; D000071857:Tumor Suppressor p53-Binding Protein 1
chemical_list: C468783:H2AX protein, human; D006657:Histones; D047908:Intracellular Signaling Peptides and Proteins; C498301:TP53BP1 protein, human; D000071857:Tumor Suppressor p53-Binding Protein 1; D006861:Hydrogen Peroxide; C576324:ATM protein, human; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins
keywords: DNA; damage; histones; oxygen; reactive