Phorbol ester exposure activates an AP-1-mediated increase in ERCC-1 messenger RNA expression in human ovarian tumor cells.
ERCC-1 is an essential gene in the nucleotide excision repair pathway, and may be essential for life. However, the mechanism of transcriptional activation and regulation of ERCC-1 gene expression is unclear. We therefore investigated the effect of the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate (TPA) on the expression of the ERCC-1 gene in A2780/CP70 human ovarian carcinoma cells. TPA induced a four- to sixfold increase in steady-state ERCC-1 messenger RNA (mRNA) levels that was time- and concentration-dependent. Nuclear run-on experiments demonstrated that the rate of transcription of ERCC-1 was approximately 2.8-fold higher in TPA-treated cells than in the controls. TPA stimulation of A2780/CP70 cells also resulted in a rapid but transient induction of c-jun and c-fos as determined by Northern and Western blot analyses, which peaked about 2 h before the peak in ERCC-1 expression. Electrophoretic mobility shift assays of nuclear extracts from TPA-treated cells revealed an increase in DNA-binding activity specific for the AP-1-like binding site in the 5'-flanking region of ERCC-1. c-Jun and c-Fos proteins were confirmed to be the components of the activated AP-1 complex by supershift analysis. The increase in AP-1 activity occurs immediately before the increase in ERCC-1 transcription. The increase in AP-1 DNA-binding activity and the increase in ERCC-1 mRNA expression were prevented by pretreatment with cycloheximide. These data suggest that AP-1 may contribute to the upregulation of ERCC-1 in response to TPA in human ovarian cancer cells.
Li
Q
Q
Developmental Therapeutics Department, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Zhang
L
L
Tsang
B
B
Gardner
K
K
Bostick-Bruton
F
F
Reed
E
E
D000546:Amanitins / Q000494:pharmacology; D001665:Binding Sites; D002273:Carcinogens / Q000494:pharmacology*; D003513:Cycloheximide / Q000494:pharmacology; D004260:DNA Repair / Q000187:drug effects*; D004268:DNA-Binding Proteins; D004720:Endonucleases; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D009363:Neoplasm Proteins / Q000235:genetics*; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010051:Ovarian Neoplasms / Q000378:metabolism / Q000473:pathology*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000235:genetics; D012334:RNA, Neoplasm / Q000096:biosynthesis* / Q000235:genetics; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D018808:Transcription Factor AP-1 / Q000502:physiology*; D014158:Transcription, Genetic / Q000187:drug effects*; D014407:Tumor Cells, Cultured
D000546:Amanitins; D002273:Carcinogens; D004268:DNA-Binding Proteins; D009363:Neoplasm Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D011500:Protein Synthesis Inhibitors; D011506:Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D018808:Transcription Factor AP-1; D003513:Cycloheximide; C071447:ERCC1 protein, human; D004720:Endonucleases; D013755:Tetradecanoylphorbol Acetate