MPF localization is controlled by nuclear export.
In eukaryotes, mitosis is initiated by M phase promoting factor (MPF), composed of B-type cyclins and their partner protein kinase, CDK1. In animal cells, MPF is cytoplasmic in interphase and is translocated into the nucleus after mitosis has begun, after which it associates with the mitotic apparatus until the cyclins are degraded in anaphase. We have used a fusion protein between human cyclin B1 and green fluorescent protein (GFP) to study this dynamic behaviour in real time, in living cells. We found that when we injected cyclin B1-GFP, or cyclin B1-GFP bound to CDK1 (i.e. MPF), into interphase nuclei it is rapidly exported into the cytoplasm. Cyclin B1 nuclear export is blocked by leptomycin B, an inhibitor of the recently identified export factor, exportin 1 (CRM1). The nuclear export of MPF is mediated by a nuclear export sequence in cyclin B1, and an export-defective cyclin B1 accumulates in interphase nuclei. Therefore, during interphase MPF constantly shuttles between the nucleus and the cytoplasm, but the bulk of MPF is retained in the cytoplasm by rapid nuclear export. We found that a cyclin mutant with a defective nuclear export signal does not enhance the premature mitosis caused by interfering with the regulatory phosphorylation of CDK1, but is more sensitive to inhibition by the Wee1 kinase.
Wellcome/CRC Institute and Department of Zoology, Tennis Court Road, Cambridge, CB2 1QR, UK.
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Hagting
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Karlsson
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Clute
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Jackman
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Pines
D001692:Biological Transport; D015415:Biomarkers; D016203:CDC2 Protein Kinase / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000037:antagonists & inhibitors; D018797:Cell Cycle Proteins / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism*; D019926:Cyclin B / Q000235:genetics / Q000378:metabolism*; D056744:Cyclin B1; D003593:Cytoplasm / Q000378:metabolism; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D049452:Green Fluorescent Proteins; D006801:Humans; D007399:Interphase; D028884:Karyopherins; D008164:Luminescent Proteins / Q000235:genetics; D008938:Mitosis / Q000502:physiology; D009687:Nuclear Proteins; D017354:Point Mutation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D020687:cdc25 Phosphatases; D000097606:Exportin 1 Protein
D015415:Biomarkers; D016203:CDC2 Protein Kinase; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D019926:Cyclin B; D056744:Cyclin B1; D005231:Fatty Acids, Unsaturated; D049452:Green Fluorescent Proteins; D028884:Karyopherins; D008164:Luminescent Proteins; D009687:Nuclear Proteins; D011505:Protein-Tyrosine Kinases; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D020687:cdc25 Phosphatases; D000097606:Exportin 1 Protein; C534699:CCNB1 protein, human; C499349:WEE1 protein, human; C517009:CDC25C protein, human; C038753:leptomycin B