Mutation of a CK2 phosphorylation site in cdc25C impairs importin alpha/beta binding and results in cytoplasmic retention.
cdc25C is a phosphatase, which activates the mitosis-promoting factor cyclin B1/cdc2 by dephosphorylation, and thus triggers G(2)/M transition. The activity of cdc25C itself is controlled by phosphorylation of certain amino-acid residues, which among other things determines the subcellular localization of the enzyme. Here, we describe a new phosphorylation site at threonine 236 of cdc25C, which is phosphorylated by protein kinase CK2. This phosphorylation site is located near the nuclear localization signal (amino acids 239-245). We demonstrate that cdc25C interacts with importin beta and the importin alpha/beta heterodimer but not with importin alpha. We further found that a cdc25C phosphorylation mutant where threonine 236 was replaced by aspartic acid as well as cdc25C phosphorylated by CK2 binds importin beta or the importin alpha/beta heterodimer less efficiently than wild type or the corresponding alanine mutant. Furthermore, the cdc25C(T236D) shows a retarded uptake into the nucleus in a cell import assay. Inhibition of protein kinase CK2 enzyme activity in vivo resulted in an enhanced nuclear localization of cdc25C. Thus, phosphorylation of cdc25C at threonine 236 is an important signal for the retention of cdc25C in the cytoplasm.
Schwindling
Sandra L
SL
Universität des Saarlandes, Medizinische Biochemie und Molekularbiologie, Gebäude 44, D-66421 Homburg, Germany.
Noll
Andreas
A
Montenarh
Mathias
M
Götz
Claudia
C
D000595:Amino Acid Sequence; D047390:Casein Kinase II; D002453:Cell Cycle / Q000235:genetics / Q000502:physiology; D018797:Cell Cycle Proteins / Q000235:genetics* / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D013912:Threonine / Q000378:metabolism; D028901:alpha Karyopherins / Q000378:metabolism*; D028961:beta Karyopherins / Q000378:metabolism*; D020687:cdc25 Phosphatases / Q000235:genetics* / Q000378:metabolism
D018797:Cell Cycle Proteins; D028901:alpha Karyopherins; D028961:beta Karyopherins; D013912:Threonine; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases