title: Cyclin D1 overexpression perturbs DNA replication and induces replication-associated DNA double-strand breaks in acquired radioresistant cells.
abstract: Fractionated radiotherapy (RT) is widely used in cancer treatment, because it preserves normal tissues. However, repopulation of radioresistant tumors during fractionated RT limits the efficacy of RT. We recently demonstrated that a moderate level of long-term fractionated radiation confers acquired radioresistance to tumor cells, which is caused by DNA-PK/AKT/GSK3β-mediated cyclin D1 overexpression. The resulting cyclin D1 overexpression leads to forced progression of the cell cycle to S-phase, concomitant with induction of DNA double-strand breaks (DSBs). In this study, we investigated the molecular mechanisms underlying cyclin D1 overexpression-induced DSBs during DNA replication in acquired radioresistant cells. DNA fiber data demonstrated that replication forks progressed slowly in acquired radioresistant cells compared with corresponding parental cells in HepG2 and HeLa cell lines. Slowly progressing replication forks were also observed in HepG2 and HeLa cells that overexpressed a nondegradable cyclin D1 mutant. We also found that knockdown of Mus81 endonuclease, which is responsible for resolving aberrant replication forks, suppressed DSB formation in acquired radioresistant cells. Consequently, Mus81 created DSBs to remove aberrant replication forks in response to replication perturbation triggered by cyclin D1 overexpression. After treating cells with a specific inhibitor for DNA-PK or ATM, apoptosis rates increased in acquired radioresistant cells but not in parental cells by inhibiting the DNA damage response to cyclin D1-mediated DSBs. This suggested that these inhibitors might eradicate acquired radioresistant cells and improve fractionated RT outcomes.
authors: Department of Environmental Health, National Institute of Public Health, Saitama, Japan. tsimura@niph.go.jp
authors: Tsutomu
authors: T
authors: Shimura
authors: Yasushi
authors: Y
authors: Ochiai
authors: Naoto
authors: N
authors: Noma
authors: Toshiyuki
authors: T
authors: Oikawa
authors: Yui
authors: Y
authors: Sano
authors: Manabu
authors: M
authors: Fukumoto
mesh_terms: D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D016923:Cell Death; D020552:Comet Assay; D019938:Cyclin D1 / Q000378:metabolism*; D051358:Cyclin-Dependent Kinase 4 / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D004261:DNA Replication; D051747:DNA-Activated Protein Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D004720:Endonucleases / Q000378:metabolism; D006367:HeLa Cells; D056945:Hep G2 Cells; D006801:Humans; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011836:Radiation Tolerance; D025521:Tumor Suppressor Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: C530104:CCND1 protein, human; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D025521:Tumor Suppressor Proteins; D019938:Cyclin D1; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D051747:DNA-Activated Protein Kinase; D017346:Protein Serine-Threonine Kinases; D051358:Cyclin-Dependent Kinase 4; D004720:Endonucleases; C498772:MUS81 protein, human
keywords: DSBs; Mus81; Perturbation of DNA replication; cyclin D1; radioresistance
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