Transactivation of the human cdc2 promoter by adenovirus E1A. E1A induces the expression and assembly of a heteromeric complex consisting of the CCAAT box binding factor, CBF/NF-Y, and a 110-kDa DNA-binding protein.
Cyclin-dependent kinases (CDKs) play an important role in the eukaryotic cell cycle progression. Cdc2 (CDK1) is expressed in late G(1)/S phase and required for G(2) to M phase transition in higher eukaryotes. The oncoproteins, SV40 large T antigen and adenovirus E1A, induce a 110-kDa protein which specifically recognizes the two inverted CCAAT motifs of the cdc2 promoter in cycling cells and plays an essential role in transactivation of the human cdc2 promoter. Since these CCAAT motifs also conform to the consensus binding sites for the ubiquitous heterotrimeric transcription factor, CBF/NF-Y, the role of CBF/NF-Y in the transactivation of the cdc2 promoter was examined in this study. Our results indicate that CBF/NF-Y and the 110-kDa protein interact with the CCAAT box motif to form a heteromeric complex. However, mutagenesis of the pentanucleotide CCAAT motif or in the presence of urea greater than 2.5 M, no heteromeric complex was formed. In contrast, the 110-kDa protein could still bind the mutant CCAAT motif or with the wild type motif in the presence of 2.5 M urea. Furthermore, E1A.12S induced the gene expression of all three subunits of CBF/NF-Y. Coexpression of E1A and a dominant negative mutant NF-YA subunit significantly reduced the E1A-mediated transactivation of the cdc2 promoter in a dose-dependent manner. These results support the conclusion that E1A protein mediates optimal transactivation of the human cdc2 promoter by inducing the expression and assembly of a heteromeric complex consisting of the 110-kDa protein and the CBF/NF-Y which interacts with the two CCAAT motifs of the cdc2 promoter.
Kao
C Y
CY
Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas 66160-7421, USA.
Tanimoto
A
A
Arima
N
N
Sasaguri
Y
Y
Padmanabhan
R
R
D017500:Adenovirus E1A Proteins / Q000235:genetics*; D001483:Base Sequence; D001665:Binding Sites; D001704:Biopolymers; D022762:CCAAT-Enhancer-Binding Proteins; D016203:CDC2 Protein Kinase / Q000235:genetics*; D002460:Cell Line; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation
D017500:Adenovirus E1A Proteins; D001704:Biopolymers; D022762:CCAAT-Enhancer-Binding Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D014157:Transcription Factors; D016203:CDC2 Protein Kinase