The cell cycle checkpoint kinase Chk2 is a negative regulator of mitotic catastrophe.
Fusion between nonsynchronized cells leads to the formation of heterokarya which transiently activate Cyclin-dependent kinase 1 (Cdk1)/cyclin B1 and enter the prophase of the cell cycle, where they arrest due to a loss of Cdk1/cyclin B1 activity, activate p53, disorganize centrosomes, and undergo apoptosis. Here, we show that the down regulation of Cdk1/cyclin B is secondary to the activation of the DNA structure checkpoint kinase Chk2. Thus, syncytia generated by the fusion of asynchronous HeLa cells contain elevated levels of active Chk2 but not Chk1. Chk2 bearing the activating phosphorylation on threonine-68 accumulates in BRCA1 nuclear bodies when the cells arrest at the G2/M boundary. Inhibition of Chk2 by transfection of a dominant-negative Chk2 mutant or a chemical inhibitor, debromohymenialdesine, stabilizes centrosomes, maintains high cyclin B1 levels, and allows for a prolonged activation of Cdk1. Under these conditions, multinuclear HeLa syncytia do not arrest at the G2/M boundary and rather enter mitotis and subsequently die during the metaphase of the cell cycle. This mitotic catastrophe is associated with the activation of the pro-apoptotic caspase-3. Inhibition of caspases allows the cells to go beyond the metaphase arrest, indicating that apoptosis is responsible for cell death by mitotic catastrophe. In another, completely different model of mitotic catastrophe, namely 14.3.3 sigma-deficient HCT116 colon carcinoma cells treated with doxorubicin, Chk2 activation was also found to be deficient as compared to 14.3.3 sigma-sufficient controls. Inhibition of Chk2 again facilitated the induction of mitotic catastrophe in HCT116 wild-type cells. In conclusion, a conflict in cell cycle progression or DNA damage can lead to mitotic catastrophe, provided that the checkpoint kinase Chk2 is inhibited. Inhibition of Chk2 thus can sensitize proliferating cells to chemotherapy-induced apoptosis.
Castedo
Maria
M
CNRS-UMR 8125, Institut Gustave Roussy, 39 rue Camille-Desmoulins, F-94805 Villejuif, France.
Perfettini
Jean-Luc
JL
Roumier
Thomas
T
Yakushijin
Kenichi
K
Horne
David
D
Medema
René
R
Kroemer
Guido
G
D048948:14-3-3 Proteins; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D014408:Biomarkers, Tumor / Q000172:deficiency; D015704:CD4 Antigens / Q000235:genetics; D016203:CDC2 Protein Kinase / Q000096:biosynthesis / Q000235:genetics; D002459:Cell Fusion; D045744:Cell Line, Tumor / Q000187:drug effects; D018385:Centrosome / Q000648:ultrastructure; D064447:Checkpoint Kinase 2; D003110:Colonic Neoplasms / Q000473:pathology; D019926:Cyclin B / Q000096:biosynthesis / Q000235:genetics; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004249:DNA Damage; D004317:Doxorubicin / Q000494:pharmacology; D005092:Exonucleases / Q000172:deficiency; D005095:Exoribonucleases; D016195:G2 Phase; D005786:Gene Expression Regulation; D018816:Genes, cdc; D015752:Genes, env; D015726:Giant Cells / Q000166:cytology / Q000201:enzymology; D006367:HeLa Cells / Q000166:cytology / Q000201:enzymology; D006801:Humans; D008938:Mitosis; D009363:Neoplasm Proteins / Q000172:deficiency; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000502:physiology
D048948:14-3-3 Proteins; D000590:Amino Acid Chloromethyl Ketones; D000903:Antibiotics, Antineoplastic; D014408:Biomarkers, Tumor; D015704:CD4 Antigens; D019926:Cyclin B; D015853:Cysteine Proteinase Inhibitors; D009363:Neoplasm Proteins; D016159:Tumor Suppressor Protein p53; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D004317:Doxorubicin; D064447:Checkpoint Kinase 2; C578634:CHEK2 protein, human; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase; D005092:Exonucleases; D005095:Exoribonucleases; C413442:SFN protein, human