Arsenite-induced mitotic death involves stress response and is independent of tubulin polymerization.
Arsenite, a known mitotic disruptor, causes cell cycle arrest and cell death at anaphase. The mechanism causing mitotic arrest is highly disputed. We compared arsenite to the spindle poisons nocodazole and paclitaxel. Immunofluorescence analysis of alpha-tubulin in interphase cells demonstrated that, while nocodazole and paclitaxel disrupt microtubule polymerization through destabilization and hyperpolymerization, respectively, microtubules in arsenite-treated cells remain comparable to untreated cells even at supra-therapeutic concentrations. Immunofluorescence analysis of alpha-tubulin in mitotic cells showed spindle formation in arsenite- and paclitaxel-treated cells but not in nocodazole-treated cells. Spindle formation in arsenite-treated cells appeared irregular and multi-polar. gamma-tubulin staining showed that cells treated with nocodazole and therapeutic concentrations of paclitaxel contained two centrosomes. In contrast, most arsenite-treated mitotic cells contained more than two centrosomes, similar to centrosome abnormalities induced by heat shock. Of the three drugs tested, only arsenite treatment increased expression of the inducible isoform of heat shock protein 70 (HSP70i). HSP70 and HSP90 proteins are intimately involved in centrosome regulation and mitotic spindle formation. HSP90 inhibitor 17-DMAG sensitized cells to arsenite treatment and increased arsenite-induced centrosome abnormalities. Combined treatment of 17-DMAG and arsenite resulted in a supra-additive effect on viability, mitotic arrest, and centrosome abnormalities. Thus, arsenite-induced abnormal centrosome amplification and subsequent mitotic arrest is independent of effects on tubulin polymerization and may be due to specific stresses that are protected against by HSP90 and HSP70.
Taylor
B Frazier
BF
Department of Pharmacology and Toxicology, Center for Environmental Genomics and Integrative Biology, Center for Genetics and Molecular Medicine, James Graham Brown Cancer Center, University of Louisville, Louisville, KY 40202, USA.
McNeely
Samuel C
SC
Miller
Heather L
HL
States
J Christopher
JC
D000970:Antineoplastic Agents / Q000633:toxicity; D000972:Antineoplastic Agents, Phytogenic / Q000633:toxicity; D017209:Apoptosis / Q000187:drug effects; D018053:Arsenites / Q000633:toxicity*; D015153:Blotting, Western; D016923:Cell Death / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000648:ultrastructure; D002470:Cell Survival / Q000187:drug effects; D018385:Centrosome / Q000187:drug effects / Q000648:ultrastructure; D005334:Fever / Q000473:pathology; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006360:Heat-Shock Proteins / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D008870:Microtubules / Q000187:drug effects / Q000648:ultrastructure; D008938:Mitosis / Q000187:drug effects*; D015739:Nocodazole / Q000633:toxicity; D017239:Paclitaxel / Q000633:toxicity; D008941:Spindle Apparatus / Q000187:drug effects / Q000648:ultrastructure; D013312:Stress, Physiological / Q000473:pathology*; D014404:Tubulin / Q000096:biosynthesis*
D000970:Antineoplastic Agents; D000972:Antineoplastic Agents, Phytogenic; D018053:Arsenites; D006360:Heat-Shock Proteins; D014404:Tubulin; C015001:arsenite; D017239:Paclitaxel; D015739:Nocodazole