Radiation-induced double-strand breaks require ATM but not Artemis for homologous recombination during S-phase.
Double-strand breaks (DSBs) are repaired by two distinct pathways, non-homologous end joining (NHEJ) and homologous recombination (HR). The endonuclease Artemis and the PIK kinase Ataxia-Telangiectasia Mutated (ATM), mutated in prominent human radiosensitivity syndromes, are essential for repairing a subset of DSBs via NHEJ in G1 and HR in G2. Both proteins have been implicated in DNA end resection, a mandatory step preceding homology search and strand pairing in HR. Here, we show that during S-phase Artemis but not ATM is dispensable for HR of radiation-induced DSBs. In replicating AT cells, numerous Rad51 foci form gradually, indicating a Rad51 recruitment process that is independent of ATM-mediated end resection. Those DSBs decorated with Rad51 persisted through S- and G2-phase indicating incomplete HR resulting in unrepaired DSBs and a pronounced G2 arrest. We demonstrate that in AT cells loading of Rad51 depends on functional ATR/Chk1. The ATR-dependent checkpoint response is most likely activated when the replication fork encounters radiation-induced single-strand breaks leading to generation of long stretches of single-stranded DNA. Together, these results provide new insight into the role of ATM for initiation and completion of HR during S- and G2-phase. The DSB repair defect during S-phase significantly contributes to the radiosensitivity of AT cells.
Institute of Radiobiology and Molecular Radiation Oncology, Philipps-University of Marburg, Baldingerstr. 35032 Marburg, Germany.
Sabrina
S
Köcher
Thorsten
T
Rieckmann
Gabor
G
Rohaly
Wael Y
WY
Mansour
Ekkehard
E
Dikomey
Irena
I
Dornreiter
Jochen
J
Dahm-Daphi
D064007:Ataxia Telangiectasia Mutated Proteins; D002453:Cell Cycle / Q000235:genetics / Q000528:radiation effects; D018797:Cell Cycle Proteins / Q000502:physiology*; D002460:Cell Line; D053903:DNA Breaks, Double-Stranded; D004268:DNA-Binding Proteins / Q000502:physiology*; D004720:Endonucleases; D006801:Humans; D009687:Nuclear Proteins / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D051135:Rad51 Recombinase / Q000032:analysis; D011836:Radiation Tolerance; D059767:Recombinational DNA Repair; D016196:S Phase / Q000235:genetics* / Q000528:radiation effects; D025521:Tumor Suppressor Proteins / Q000502:physiology*
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D025521:Tumor Suppressor Proteins; C576324:ATM protein, human; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D051135:Rad51 Recombinase; C426344:DCLRE1C protein, human; D004720:Endonucleases