Activation-dependent and activation-independent localisation of calmodulin to the mitotic apparatus during the first cell cycle of the Lytechinus pictus embryo.
We have used confocal microscopy and a fluorescent calmodulin probe to examine the mechanism of localisation of calmodulin during the first cell cycle of the sea urchin zygote. Using fluorescein-calmodulin, calmodulin can be observed within the nucleus and interphase astral microtubule arrays as cells approach mitosis. During mitosis, calmodulin redistributes to the mitotic apparatus and to condensed chromosomes. Quantitative analysis with reference to a control dye (fluorescein-dextran) shows that the distribution of calmodulin is specific. We used a competitive inhibitor of calcium-dependent calmodulin binding (Trp-peptide; Torok & Trentham (1994) Biochemistry 33, 12807-20) to test whether the cell cycle localisation of calmodulin was due to its binding to targets on activation. The Trp-peptide eliminates localisation of calmodulin within the nucleus. However, microtubule localisation persists in the presence of the Trp-peptide. These data show that calmodulin can localise by calcium (and hence activation)-dependent as well as calcium-independent mechanisms. This suggests that distinct mechanisms of localisation may be involved in the regulation of the differential functions of calmodulin, at least during the cell cycle.
Department of Physiology, University College London, UK.
M
M
Wilding
K
K
Torok
M
M
Whitaker
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002453:Cell Cycle / Q000235:genetics*; D002467:Cell Nucleus / Q000378:metabolism; D003911:Dextrans / Q000378:metabolism; D005306:Fertilization; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D008845:Microinjections; D018613:Microscopy, Confocal / Q000379:methods; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000378:metabolism; D008938:Mitosis / Q000502:physiology*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D012617:Sea Urchins / Q000196:embryology / Q000378:metabolism*; D015053:Zygote / Q000378:metabolism*
D002147:Calmodulin; D003911:Dextrans; D005452:Fluoresceins; D005456:Fluorescent Dyes; D010446:Peptide Fragments; C052526:fluorescein-dextran; D002118:Calcium