title: Three independent mechanisms for arrest in G2 after ionizing radiation.
abstract: Cell cycle checkpoints ensure that eukaryotic cells do not enter mitosis after ionizing irradiation (IR). The G(2)-arrest after IR is the result of activation of multiple signalling pathways, the contributions of which vary with time after irradiation. We have studied the time evolution of the IR-induced G(2)-arrest in human B-lymphocyte cancer cell lines, as well as the molecular mechanisms responsible for the arrest. Cells that were in G(2) phase at the time of irradiation experienced a transient arrest that blocked entry into mitosis at 0-2 hours after IR (0.5 or 4 Gy). Activation of ATM and CHEK2 occurred at the same time as this early arrest and was, like the arrest, abrogated by the ATM-inhibitor KU-55933. A late, permanent and ATM-independent arrest (≥6 hours after IR) of cells that were in G(2)/S/G(1) at the time of irradiation (4 Gy) was inactivated by caffeine. This late G(2)-arrest could not be explained by down-regulation of genes with functions in G(2)/mitosis (e.g. PLK1, CCNB1/2), since the down-regulation was transient and not accompanied by reduced protein levels. However, the persistent phosphorylation of CHEK1 after 4 Gy suggested a role for CHEK1 in the late arrest, consistent with the abrogation of the arrest in CHEK1-depleted cells. TP53 was not necessary for the late G(2)-arrest, but mediated an intermediate arrest (2-10 hours after IR) independently of ATM and CHEK1. In conclusion, the IR-induced arrest in G(2) is mediated by ATM immediately after irradiation, with TP53 for independent and transient back-up, while CHEK1 is necessary for the late arrest.
authors.affiliation: Department of Radiation Biology, Institute for Cancer Research, The Norwegian Radium Hospital, Oslo, Norway.
authors.forename: Kirsti Solberg
authors.initials: KS
authors.lastname: Landsverk
authors.forename: Sebastian
authors.initials: S
authors.lastname: Patzke
authors.forename: Idun Dale
authors.initials: ID
authors.lastname: Rein
authors.forename: Caroline
authors.initials: C
authors.lastname: Stokke
authors.forename: Heidi
authors.initials: H
authors.lastname: Lyng
authors.forename: Paula M
authors.initials: PM
authors.lastname: De Angelis
authors.forename: Trond
authors.initials: T
authors.lastname: Stokke
mesh_terms: D064007:Ataxia Telangiectasia Mutated Proteins; D002110:Caffeine / Q000494:pharmacology; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1; D064447:Checkpoint Kinase 2; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D016195:G2 Phase / Q000528:radiation effects*; D006801:Humans; D008938:Mitosis; D009025:Morpholines / Q000494:pharmacology; D010766:Phosphorylation; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011753:Pyrones / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D011839:Radiation, Ionizing; D015398:Signal Transduction; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: C495818:2-morpholin-4-yl-6-thianthren-1-yl-pyran-4-one; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D009025:Morpholines; D011753:Pyrones; D034741:RNA, Small Interfering; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D002110:Caffeine; D011494:Protein Kinases; D064447:Checkpoint Kinase 2; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C000605867:CHEK1 protein, human; C578634:CHEK2 protein, human; D000071877:Checkpoint Kinase 1; D017346:Protein Serine-Threonine Kinases