Ca2+ signal blockers can inhibit M/A transition in mammalian cells by interfering with the spindle checkpoint.
A key step in mitosis is the sister-chromatid separation at the metaphase-anaphase (M/A) transition. Several earlier studies had suggested that Ca(2+) signal is involved in regulating this process in somatic cells. The detailed mechanisms, however, are not yet well understood. In this study, we used the GFP-gene fusion method and a living-cell imaging technique to examine the effects of suppressing cytosolic Ca(2+) level on the mitotic process in HeLa and PtK2 cells. We observed that application of the Ca(2+) chelator BAPTA/AM can block or severely delay the M/A transition. This blockage was caused by a failure in activating the anaphase-promoting complex (APC), since both cyclin B and securin could not be degraded under this situation. Furthermore, using YFP-labeled tubulin, we found that the mitotic spindle structure in most of the BAPTA-treated cells gradually deformed with time. Other Ca(2+) signal blockers, such as heparin, also produced a similar effect. These results suggest that one pathway for the blockage of M/A transition by suppressing cytosolic Ca(2+) level is due to its interference with the mitotic spindle checkpoint.
Xu
Naihan
N
Department of Biology, The Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, PR China.
Luo
Kathy Q
KQ
Chang
Donald C
DC
D000705:Anaphase / Q000502:physiology*; D064173:Anaphase-Promoting Complex-Cyclosome; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D002460:Cell Line; D002614:Chelating Agents / Q000494:pharmacology; D019926:Cyclin B / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D008025:Ligases / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism; D008677:Metaphase / Q000502:physiology*; D009363:Neoplasm Proteins / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D064249:Securin; D008941:Spindle Apparatus / Q000502:physiology*; D014404:Tubulin / Q000378:metabolism; D043743:Ubiquitin-Protein Ligase Complexes
D002614:Chelating Agents; D019926:Cyclin B; D008164:Luminescent Proteins; D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins; D064249:Securin; D014404:Tubulin; C578002:pituitary tumor-transforming protein 1, human; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D008025:Ligases; D002118:Calcium