Mitogen-induced expression of the fibroblast growth factor-binding protein is transcriptionally repressed through a non-canonical E-box element.
The fibroblast growth factor-binding protein (FGF-BP) stimulates FGF-2-mediated angiogenesis and is thought to play an important role in the progression of squamous cell, colon, and breast carcinomas. 12-O-Tetradecanoylphorbol-13-acetate (TPA) induction of the FGF-BP gene occurs through transcriptional mechanisms involving Sp1, AP-1, and CCAATT/enhancer-binding protein sites in the proximal FGF-BP gene promoter. The level of TPA induction, however, is limited due to the presence of a repressor element that shows similarity to a non-canonical E-box (AACGTG). Mutation or deletion of the repressor element led to enhanced induction by TPA or epidermal growth factor in cervical squamous cell and breast carcinoma cell lines. Repression was dependent on the adjacent AP-1 site, without discernible alteration in the binding affinity or composition of AP-1. We investigated the following two possible mechanisms for E-box-mediated repression: 1) CpG methylation of the core of the E-box element, and 2) binding of a distinct protein complex to this site. Point mutation of the CpG methylation site in the E-box showed loss of repressor activity. Conversely, in vitro methylation of this site significantly reduced TPA induction. In vitro gel shift analysis revealed distinct and TPA-dependent binding of USF1 and USF2 to the repressor element that required nucleotides within the E-box. Furthermore, chromatin immunoprecipitation assay showed that USF, c-Myc, and Max proteins were associated with the FGF-BP promoter in vivo. Overall, these findings suggested that the balance between trans-activation by AP-1 and repression through the E-box is an important control mechanism for fine-tuning the angiogenic response to growth factor-activated pathways.
Harris
V K
VK
Department of Oncology, Vincent T. Lombardi Cancer Center, Georgetown University, Washington, D. C. 20007, USA.
Coticchia
C M
CM
List
H J
HJ
Wellstein
A
A
Riegel
A T
AT
D022762:CCAAT-Enhancer-Binding Proteins; D002352:Carrier Proteins / Q000235:genetics*; D002460:Cell Line; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000502:physiology; D005346:Fibroblast Growth Factors / Q000494:pharmacology; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D009687:Nuclear Proteins / Q000502:physiology; D011401:Promoter Regions, Genetic; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D012097:Repressor Proteins / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D018808:Transcription Factor AP-1 / Q000502:physiology
D022762:CCAAT-Enhancer-Binding Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D036341:Intercellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D016271:Proto-Oncogene Proteins c-myc; D012097:Repressor Proteins; D018808:Transcription Factor AP-1; C070699:FGFBP1 protein, human; D005346:Fibroblast Growth Factors; D013755:Tetradecanoylphorbol Acetate