CCAAT/enhancer-binding protein-alpha cooperates with p21 to inhibit cyclin-dependent kinase-2 activity and induces growth arrest independent of DNA binding.
CCAAT/enhancer-binding protein-alpha (C/EBP alpha) is a basic leucine zipper protein that controls transcription of genes important for liver function, white adipose tissue development, and granulocyte differentiation. In addition to its function in controlling gene expression in differentiated tissues, C/EBP alpha is also associated with an antimitotic activity. We have previously demonstrated that C/EBP alpha interacts with p21, a cyclin-dependent kinase (CDK) inhibitor, and that C/EBP alpha inhibits proliferation when expressed in several different cell types (Timchenko, N. A., Harris, T. E., Wilde, M., Bilyeu, T. A., Burgess-Beusse, B. L., Finegold, M. J., and Darlington, G. J. (1997) Mol. Cell. Biol. 17, 7353--7361). Here we define the regions of C/EBP alpha required for interaction with p21 and demonstrate that CDK2 also interacts with C/EBP alpha. We show that C/EBP alpha can cooperate with p21 to inhibit CDK2 activity in vitro. The effect of C/EBP alpha on CDK2 activity requires the p21 and CDK2 interaction sites within C/EBP alpha. C/EBP alpha mutants incapable of inhibiting CDK2 activity in vitro do not inhibit proliferation in cultured cells. However, C/EBP alpha mutants defective in DNA binding inhibit proliferation as effectively as the wild-type protein. These findings show that C/EBP alpha-mediated growth arrest occurs through protein interactions and is independent of its transcriptional activity.
Harris
T E
TE
Cell and Molecular Biology Program, Baylor College of Medicine, Houston, Texas 77030, USA.
Albrecht
J H
JH
Nakanishi
M
M
Darlington
G J
GJ
D019943:Amino Acid Substitution; D000818:Animals; D022762:CCAAT-Enhancer-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D042846:CDC2-CDC28 Kinases; D002453:Cell Cycle / Q000502:physiology*; D002455:Cell Division; D002460:Cell Line; D019927:Cyclin E / Q000378:metabolism*; D051357:Cyclin-Dependent Kinase 2; D050759:Cyclin-Dependent Kinase Inhibitor p21; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors* / Q000302:isolation & purification / Q000378:metabolism; D016213:Cyclins / Q000302:isolation & purification / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology; D006801:Humans; D007700:Kinetics; D051379:Mice; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D017354:Point Mutation; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism; D017384:Sequence Deletion; D018411:Spodoptera; D014162:Transfection
D022762:CCAAT-Enhancer-Binding Proteins; C493956:CDKN1A protein, human; C493957:Cdkn1a protein, mouse; D019927:Cyclin E; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; C495893:Cdk2 protein, mouse; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases