Hyperphosphorylation of the N-terminal domain of Cdc25 regulates activity toward cyclin B1/Cdc2 but not cyclin A/Cdk2.
Cdc25 regulates entry into mitosis by regulating the activation of cyclin B/cdc2. In humans, at least two cdc25 isoforms have roles in controlling the G2/M transition. Here we show, using bacterially expressed recombinant proteins, that two cdc25B splice variants, cdc25B2 and cdc25B3, are capable of activating cyclin A/cdk2 and cyclin B/cdc2, but that mitotic hyperphosphorylation of these proteins increases their activity toward only cyclin B1/cdc2. Cdc25C has only very low activity in its unphosphorylated form, and following hyperphosphorylation it will efficiently catalyze the activation of only cyclin B/cdc2. This was reflected by the in vivo activity of the immunoprecipitated cdc25B and cdc25C from interphase and mitotic HeLa cells. The increased activity of the hyperphosphorylated cdc25s toward cyclin B1/cdc2 was in large part due to increased binding of this substrate. The substrate specificity, activities, and timing of the hyperphosphorylation of cdc25B and cdc25C during G2 and M suggest that these two mitotic cdc25 isoforms are activated by different kinases and perform different functions during progression through G2 into mitosis.
Gabrielli
B G
BG
Queensland Cancer Fund Research Laboratories and Joint Oncology Program, Queensland Institute of Medical Research, P. O. Royal Brisbane Hospital, Queensland, Australia 4029. brianG@qimr.edu.au
Clark
J M
JM
McCormack
A K
AK
Ellem
K A
KA
D016203:CDC2 Protein Kinase / Q000378:metabolism*; D042846:CDC2-CDC28 Kinases; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D019925:Cyclin A / Q000378:metabolism*; D019926:Cyclin B / Q000378:metabolism*; D056744:Cyclin B1; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007399:Interphase; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020687:cdc25 Phosphatases
C534699:CCNB1 protein, human; D018797:Cell Cycle Proteins; D019925:Cyclin A; D019926:Cyclin B; D056744:Cyclin B1; D011993:Recombinant Fusion Proteins; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases; D010749:Phosphoprotein Phosphatases; C517004:CDC25A protein, human; C517008:CDC25B protein, human; D017027:Protein Tyrosine Phosphatases; D020687:cdc25 Phosphatases; D019204:GTP-Binding Proteins