Cdc25A localisation and shuttling: characterisation of sequences mediating nuclear export and import.
The Cdc25 phosphatases play crucial roles in cell cycle progression by removing inhibitory phosphates from tyrosine and threonine residues of cyclin-dependent kinases. Cdc25A is an important regulator of the G1/S transition but functions also in the mitotic phase of the human cell cycle. In this paper, we investigate the sub-cellular localisation of exogenously expressed Cdc25A. We show that YFP-Cdc25A is localised both in the nucleus and the cytoplasm of HeLa cells and untransformed fibroblasts. Cell fusion assays and fluorescence loss in photobleaching (FLIP) assays reveal that the localisation is dynamic and the protein shuttles between the nucleus and the cytoplasm. We demonstrate that nuclear export of Cdc25A is partly mediated by an N-terminal nuclear export sequence (NES), in a manner not sensitive to the Exportin 1-inhibitor leptomycin B. A nuclear localisation signal (NLS) is also characterised, mutation of which leads to cytoplasmic localisation of Cdc25A. Our results imply that the Cdc25A phosphatase may interact with substrates and regulators both in the nucleus and the cytoplasm.
Källström
Helena
H
Department of Cell and Molecular Biology, Karolinska Institutet, 171 77 Stockholm, Sweden.
Lindqvist
Arne
A
Pospisil
Vitek
V
Lundgren
Andreas
A
Rosenthal
Christina Karlsson
CK
D021581:Active Transport, Cell Nucleus / Q000502:physiology*; D000595:Amino Acid Sequence; D002467:Cell Nucleus / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D028884:Karyopherins / Q000378:metabolism; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D020687:cdc25 Phosphatases / Q000378:metabolism*; D000097606:Exportin 1 Protein
D028884:Karyopherins; D019913:Nuclear Localization Signals; D018160:Receptors, Cytoplasmic and Nuclear; D020687:cdc25 Phosphatases; D000097606:Exportin 1 Protein; C517004:CDC25A protein, human