Interaction of the CDK2-associated protein-1, p12^{DOC-1/CDK2AP1}, with its homolog, p14^DOC-1R

Abstract

Human DOC-1/CDK2AP1 gene encodes a growth suppressor protein of 12kDa (p12DOC-1/CDK2AP1). Recently, p12DOC-1/CDK2AP1has been shown to associate with cell cycle proteins including CDK2 and DNA polymerase α/primase. It negatively regulates CDK2 activities and suppresses DNA replication. Therefore, identification of other p12DOC-1/CDK2AP1interacting proteins might clarify its role in the cell cycle regulation and carcinogenesis. The purpose of this study was to identify additional p12DOC-1/CDK2AP1interacting proteins using the yeast two-hybrid system. Using human p12DOC-1/CDK2AP1as a bait in a liver cDNA library screening, cDNA clones identical to human DOC-1R transcript were identified. The interaction between p12DOC-1/CDK2AP1and p14DOC-1Rwas verified in vitro and in cells. GST pull-down assay and immunoprecipitation experiments confirmed the interaction between the two proteins. The critical region for p12DOC-1/CDK2AP1interaction with p14DOC-1Rwas defined to amino acids 20-25 by using a series of deletion mutants as baits in the yeast two-hybrid system. Our data indicated that p12DOC-1/CDK2AP1could associate with its homologous protein, p14DOC-1R. © 2004 Elsevier Inc. All rights reserved.