Bleomycin-induced over-replication involves sustained inhibition of mitotic entry through the ATM/ATR pathway.
Polyploid cells result in aneuploidy through aberrant chromosome segregation, possibly leading to tumorigenesis. Although polyploid cells are induced through over-replication by a variety of agents, including DNA-damaging drugs, the mechanisms that induce polyploidy have been hitherto unknown. Here, we show that treatment with bleomycin, a glycopeptide anticancer drug, induces over-replication at low cytotoxic doses. During bleomycin-induced over-replication, mitotic entry is inhibited through tyrosine phosphorylation of CDK1 along the ATM/ATR pathway in the early phase of treatment. Bleomycin-induced over-replication is inhibited by the inhibitors of the ATM/ATR pathway through abrogation of bleomycin-induced G2 arrest, and the ATM/ATR inhibitors promote cell death instead of over-replication. Following the phosphorylation of CDK1, the level of cyclin B1 is decreased in the late phase of treatment. Time-lapse imaging of clone cells that express a live cell marker of endogenous cyclin B1 revealed that cyclin B1 is degraded in G2-arrested cells upon bleomycin treatment. Our findings lead to a model of how the ATM/ATR pathway acts as a molecular switch for regulating cell fates, flipping between cell death via progress into mitosis, and over-replication via sustained G2 arrest upon DNA damage, where cyclin B1 degradation is an important factor for inducing over-replication.
Nakayama
Yuji
Y
Department of Molecular Cell Biology, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba, Japan. nakayama@p.chiba-u.ac.jp
Igarashi
Asae
A
Kikuchi
Ikue
I
Obata
Yuuki
Y
Fukumoto
Yasunori
Y
Yamaguchi
Naoto
N
D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D064007:Ataxia Telangiectasia Mutated Proteins; D001761:Bleomycin / Q000494:pharmacology*; D016203:CDC2 Protein Kinase / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects / Q000502:physiology; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D019926:Cyclin B / Q000378:metabolism; D056744:Cyclin B1; D004261:DNA Replication / Q000187:drug effects*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004317:Doxorubicin / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis / Q000187:drug effects*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*
D000903:Antibiotics, Antineoplastic; C534699:CCNB1 protein, human; D018797:Cell Cycle Proteins; D019926:Cyclin B; D056744:Cyclin B1; D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D025521:Tumor Suppressor Proteins; D001761:Bleomycin; D004317:Doxorubicin; C576324:ATM protein, human; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase