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 zippe r protein that controls transcription of genes important for liver function , white adipose tissue development, and granulocyte differentiation. In add ition to its function in controlling gene expression in differentiated tiss ues, C/EBP alpha is also associated with an antimitotic activity. We have p reviously demonstrated that C/EBP alpha interacts with p21, a cyclin-depend ent kinase (CDK) inhibitor, and that C/EBP alpha inhibits proliferation whe n 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 Da rlington, G. J. (1997) Mol. Cell. Biol. 17,7353-7361). Here we define the r egions of C/EBP alpha required for interaction with p21 and demonstrate tha t CDK2 also interacts with C/EBP alpha. We show that C/EBP alpha can cooper ate 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 a lpha. 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-t ype protein. These findings show that C/EBP alpha -mediated growth arrest o ccurs through protein interactions and is independent of its transcriptiona l activity.