Activation of the murine dihydrofolate reductase promoter by E2F1

A requirement for CBP recruitment

Christopher J. Fry, Angela Pearson, Erik Malinowski, Stephanie M. Bartley, Jack Greenblatt, Peggy J. Farnhamt

Research output: Contribution to journalArticle

43 Citations (Scopus)

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.

Original languageEnglish (US)
Pages (from-to)15883-15891
Number of pages9
JournalJournal of Biological Chemistry
Volume274
Issue number22
DOIs
StatePublished - May 28 1999
Externally publishedYes

Fingerprint

Tetrahydrofolate Dehydrogenase
Chemical activation
TATA-Box Binding Protein
Phenylalanine
S Phase
Transcriptional Activation
Cell Cycle
Transcription Factor TFIIH
Binding Sites
Cells
Amino Acids
Functional analysis
Trans-Activators
Phase Transition
G1 Phase
Gene Expression Regulation
Amino Acid Substitution
Gene expression
Alanine
Proteins

ASJC Scopus subject areas

  • Biochemistry

Cite this

Fry, C. J., Pearson, A., Malinowski, E., Bartley, S. M., Greenblatt, J., & Farnhamt, P. J. (1999). Activation of the murine dihydrofolate reductase promoter by E2F1: A requirement for CBP recruitment. Journal of Biological Chemistry, 274(22), 15883-15891. https://doi.org/10.1074/jbc.274.22.15883

Activation of the murine dihydrofolate reductase promoter by E2F1 : A requirement for CBP recruitment. / Fry, Christopher J.; Pearson, Angela; Malinowski, Erik; Bartley, Stephanie M.; Greenblatt, Jack; Farnhamt, Peggy J.

In: Journal of Biological Chemistry, Vol. 274, No. 22, 28.05.1999, p. 15883-15891.

Research output: Contribution to journalArticle

Fry, CJ, Pearson, A, Malinowski, E, Bartley, SM, Greenblatt, J & Farnhamt, PJ 1999, 'Activation of the murine dihydrofolate reductase promoter by E2F1: A requirement for CBP recruitment', Journal of Biological Chemistry, vol. 274, no. 22, pp. 15883-15891. https://doi.org/10.1074/jbc.274.22.15883
Fry, Christopher J. ; Pearson, Angela ; Malinowski, Erik ; Bartley, Stephanie M. ; Greenblatt, Jack ; Farnhamt, Peggy J. / Activation of the murine dihydrofolate reductase promoter by E2F1 : A requirement for CBP recruitment. In: Journal of Biological Chemistry. 1999 ; Vol. 274, No. 22. pp. 15883-15891.
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