Mitotic arrest-associated apoptosis induced by sodium arsenite in A375 melanoma cells is BUBR1-dependent.
A375 human malignant melanoma cells undergo mitotic arrest-associated apoptosis when treated with pharmacological concentrations of sodium arsenite, a chemotherapeutic for acute promyelocytic leukemia. Our previous studies indicated that decreased arsenite sensitivity correlated with reduced mitotic spindle checkpoint function and reduced expression of the checkpoint protein BUBR1. In the current study, arsenite induced securin and cyclin B stabilization, BUBR1 phosphorylation, and spindle checkpoint activation. Arsenite also increased activating cyclin dependent kinase 1 (CDK1) Thr(161) phosphorylation but decreased inhibitory Tyr15 phosphorylation. Mitotic arrest resulted in apoptosis as indicated by colocalization of mitotic phospho-Histone H3 with active caspase 3. Apoptosis was associated with BCL-2 Ser70 phosphorylation. Inhibition of CDK1 with roscovitine in arsenite-treated mitotic cells inhibited spindle checkpoint maintenance as inferred from reduced BUBR1 phosphorylation, reduced cyclin B expression, and diminution of mitotic index. Roscovitine also reduced BCL-2 Ser70 phosphorylation and protected against apoptosis, suggesting mitotic arrest caused by hyperactivation of CDK1 directly or indirectly leads to BCL-2 phosphorylation and apoptosis. In addition, suppression of BUBR1 with siRNA prevented arsenite-induced mitotic arrest and apoptosis. These findings provide insight into the mechanism of arsenic's chemotherapeutic action and indicate a functional spindle checkpoint may be required for arsenic-sensitivity.
McNeely
Samuel C
SC
Department of Pharmacology and Toxicology, Center for Environmental Genomics and Integrative Biology, Center for Genetics and Molecular Medicine and Brown Cancer Center, University of Louisville, 570 S. Preston St. Suite 221, Louisville, KY 40202, USA.
Taylor
B Frazier
BF
States
J Christopher
JC
D000970:Antineoplastic Agents / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D018053:Arsenites / Q000633:toxicity*; D015153:Blotting, Western; D016203:CDC2 Protein Kinase / Q000235:genetics / Q000502:physiology; D005455:Fluorescent Antibody Technique; D006801:Humans; D008546:Melanoma, Experimental / Q000473:pathology*; D008938:Mitosis / Q000187:drug effects*; D015739:Nocodazole / Q000494:pharmacology; D017239:Paclitaxel / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000502:physiology*; D011687:Purines / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000494:pharmacology; D000077546:Roscovitine; D017670:Sodium Compounds / Q000633:toxicity*
D000970:Antineoplastic Agents; D000972:Antineoplastic Agents, Phytogenic; D018053:Arsenites; D011687:Purines; D034741:RNA, Small Interfering; D017670:Sodium Compounds; D000077546:Roscovitine; C017947:sodium arsenite; C584277:BUB1 protein, human; C090464:Bub1 spindle checkpoint protein; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase; D017239:Paclitaxel; D015739:Nocodazole