title: Activation of the murine dihydrofolate reductase promoter by E2F1. A requirement for CBP recruitment.
abstract: The E2F family of heterodimeric transcription factors plays an important role in the regulation of gene expression at the G1/S phase transition of the mammalian cell cycle. Previously, we have demonstrated that cell cycle regulation of murine dihydrofolate reductase (dhfr) expression requires E2F-mediated activation of the dhfr promoter in S phase. To investigate the mechanism by which E2F activates an authentic E2F-regulated promoter, we precisely replaced the E2F binding site in the dhfr promoter with a Gal4 binding site. Using Gal4-E2F1 derivatives, we found that E2F1 amino acids 409-437 contain a potent core transactivation domain. Functional analysis of the E2F1 core domain demonstrated that replacement of phenylalanine residues 413, 425, and 429 with alanine reduces both transcriptional activation of the dhfr promoter and protein-protein interactions with CBP, transcription factor (TF) IIH, and TATA-binding protein (TBP). However, additional amino acid substitutions for phenylalanine 429 demonstrated a strong correlation between activation of the dhfr promoter and binding of CBP, but not TFIIH or TBP. Finally, transactivator bypass experiments indicated that direct recruitment of CBP is sufficient for activation of the dhfr promoter. Therefore, we suggest that recruitment of CBP is one mechanism by which E2F activates the dhfr promoter.
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin Medical School, Madison, Wisconsin 53706, USA.
authors.forename: C J
authors.initials: CJ
authors.lastname: Fry
authors.forename: A
authors.initials: A
authors.lastname: Pearson
authors.forename: E
authors.initials: E
authors.lastname: Malinowski
authors.forename: S M
authors.initials: SM
authors.lastname: Bartley
authors.forename: J
authors.initials: J
authors.lastname: Greenblatt
authors.forename: P J
authors.initials: PJ
authors.lastname: Farnham
mesh_terms: D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D050882:CREB-Binding Protein; D002352:Carrier Proteins; D002453:Cell Cycle; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins / Q000235:genetics; D029721:Drosophila Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D005656:Fungal Proteins / Q000235:genetics; D005786:Gene Expression Regulation; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics; D051736:Proto-Oncogene Proteins c-mdm2; D016160:Retinoblastoma Protein / Q000235:genetics; D056465:Retinoblastoma-Binding Protein 1; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D035381:TATA-Binding Protein Associated Factors; D035181:TATA-Box Binding Protein; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics*; D015534:Trans-Activators / Q000235:genetics*; D050680:Transcription Factor DP1; D035362:Transcription Factor TFIID; D051758:Transcription Factor TFIIH; D014157:Transcription Factors / Q000235:genetics*; D019473:Transcription Factors, TFII; D015533:Transcriptional Activation / Q000235:genetics; D014162:Transfection
chemical_list: C533431:Arid4a protein, mouse; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C088053:Dp transcription factor, Drosophila; D029721:Drosophila Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493754:E2f1 protein, mouse; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D016160:Retinoblastoma Protein; D056465:Retinoblastoma-Binding Protein 1; D029701:Saccharomyces cerevisiae Proteins; C490039:TAF6 protein, S cerevisiae; D035381:TATA-Binding Protein Associated Factors; D035181:TATA-Box Binding Protein; D015534:Trans-Activators; D050680:Transcription Factor DP1; D035362:Transcription Factor TFIID; D014157:Transcription Factors; D019473:Transcription Factors, TFII; D051758:Transcription Factor TFIIH; D013762:Tetrahydrofolate Dehydrogenase; D050882:CREB-Binding Protein; C494364:Crebbp protein, mouse; C497207:Mdm2 protein, mouse; D051736:Proto-Oncogene Proteins c-mdm2