Interaction of the CDK2-associated protein-1, p12(DOC-1/CDK2AP1), with its homolog, p14(DOC-1R).
Human DOC-1/CDK2AP1 gene encodes a growth suppressor protein of 12kDa (p12(DOC-1/CDK2AP1)). Recently, p12(DOC-1/CDK2AP1) has been shown to associate with cell cycle proteins including CDK2 and DNA polymerase alpha/primase. It negatively regulates CDK2 activities and suppresses DNA replication. Therefore, identification of other p12(DOC-1/CDK2AP1) interacting proteins might clarify its role in the cell cycle regulation and carcinogenesis. The purpose of this study was to identify additional p12(DOC-1/CDK2AP1) interacting proteins using the yeast two-hybrid system. Using human p12(DOC-1/CDK2AP1) as a bait in a liver cDNA library screening, cDNA clones identical to human DOC-1R transcript were identified. The interaction between p12(DOC-1/CDK2AP1) and p14(DOC-1R) was verified in vitro and in cells. GST pull-down assay and immunoprecipitation experiments confirmed the interaction between the two proteins. The critical region for p12(DOC-1/CDK2AP1)'s interaction with p14(DOC-1R) was 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 p12(DOC-1/CDK2AP1) could associate with its homologous protein, p14(DOC-1R).
Department of Oral Medicine, Faculty of Dentistry, Mahidol University, Bangkok, Thailand.
Waranun
W
Buajeeb
Xue
X
Zhang
Hiroe
H
Ohyama
David
D
Han
Rudee
R
Surarit
Yong
Y
Kim
David T W
DT
Wong
D000595:Amino Acid Sequence; D042846:CDC2-CDC28 Kinases / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D016147:Genes, Tumor Suppressor; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015513:Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D014162:Transfection; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
C096151:CDK2AP1 protein, human; C401561:CDK2AP2 protein, human; D015513:Oncogene Proteins; D011993:Recombinant Fusion Proteins; D025521:Tumor Suppressor Proteins; D042846:CDC2-CDC28 Kinases