Downregulation of cytochromes P450 in growth-stimulated rat hepatocytes: role of c-Myc induction and impaired C/EBP binding to DNA.
OBJECTIVE
Several cytochromes P450 (CYPs) are expressed in differentiated hepatocytes, but downregulated in growth-stimulated cells. We determined the signals involved in CYP downregulation by epidermal growth factor (EGF).


METHODS
Rat hepatocytes were cultured with or without diverse substances for 72 h and EGF for the last 48 h.


RESULTS
EGF increased c-myc mRNA and protein, and decreased CYP mRNAs and proteins; both effects were prevented by two agents blocking c-myc transcription (retinoic acid and DMSO) and two antisense c-myc oligomers. Despite unchanged CCAAT-enhancer binding protein alpha (C/EBPalpha) and increased C/EBPbeta levels, nuclear proteins of EGF-treated cells did not bind to a C/EBP DNA probe in a gel mobility shift assay. This binding was restored when cells were co-treated with both EGF and c-myc antisense oligomers (preventing c-Myc induction). The N-terminal c-Myc domain added to control nuclear extracts prevented C/EBP DNA binding. A monoclonal anti-c-Myc antibody co-immunoprecipitated c-Myc, C/EBPalpha and C/EBPbeta from nuclear extracts. In cells not treated with EGF, an antisense C/EBPalpha oligomer decreased CYP expression.


CONCLUSIONS
EGF overexpresses c-Myc, decreases C/EBP binding to DNA and downregulates CYPs. We suggest that c-Myc may form inactive complexes with C/EBPs, thus decreasing C/EBP-mediated CYP transactivation.
Tinel
Marina
M
Institut National de la Santé et de la Recherche Médicale (INSERM) Unité 481, Faculté de Médecine Xavier Bichat, 16 rue Henri Huchard, 75018, Paris, France.
Berson
Alain
A
Elkahwaji
Johny
J
Cresteil
Thierry
T
Beaune
Philippe
P
Pessayre
Dominique
D
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000378:metabolism*; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology; D016166:Free Radical Scavengers / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D022781:Hepatocytes / Q000166:cytology / Q000502:physiology*; D008297:Male; D016376:Oligonucleotides, Antisense; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D014212:Tretinoin / Q000494:pharmacology
D000970:Antineoplastic Agents; D022763:CCAAT-Enhancer-Binding Protein-alpha; D016166:Free Radical Scavengers; D016376:Oligonucleotides, Antisense; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D014212:Tretinoin; D004815:Epidermal Growth Factor; D003577:Cytochrome P-450 Enzyme System; D004121:Dimethyl Sulfoxide