Down-regulation of nuclear aryl hydrocarbon receptor DNA-binding and transactivation functions: requirement for a labile or inducible factor.
Aryl hydrocarbons (AHs) such as 2,3,7,8-tetrachlorodibenzo-p-dioxin and benzo[a]pyrene activate the sequence-specific DNA-binding activity of the AH receptor. In the rat hepatocyte-derived cell line LCS7, DNA-binding activity peaked after 30 min and was then down-regulated, reaching negligible levels by 2 h. Down-regulation could be blocked, and DNA-binding activity maintained at maximum for many hours by inhibiting protein or RNA synthesis, implying that down-regulation is a mediated process requiring a labile or inducible protein. CYP1A1 transcription and in vivo DNA-protein interactions at xenobiotic response elements were down-regulated in parallel with DNA-binding activity in nuclear extracts, and these changes could also be blocked by inhibitors of protein synthesis. The correlation between AH receptor DNA-binding activity, intensity of in vivo footprints at xenobiotic response elements, and CYP1A1 transcription rate implies that down-regulation of AH receptor DNA-binding activity is important in regulating CYP1A1 transcription and that receptor is required continuously to maintain transcription. This correlation extends to the murine hepatoma cell line Hepa-1c1c7, in which slower kinetics of activation and down-regulation of CYP1A1 transcription paralleled slower activation and down-regulation of AH receptor DNA-binding activity. The difference in kinetics between cell lines also implies that AH receptor DNA-binding activity is modulated by a mechanism that may be influenced by cell-specific regulatory pathways. The above observations in conjunction with mixing experiments and comparisons of cytoplasmic and nuclear extracts indicate that down-regulation of AH receptor DNA-binding activity is probably due either to degradation or to conversion of the receptor to form that is inactive in both DNA binding and transactivation.
Reick
M
M
Molecular Biology Laboratory, Maharishi International University, Fairfield, Iowa 52557-1078, USA.
Robertson
R W
RW
Pasco
D S
DS
Fagan
J B
JB
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015536:Down-Regulation; D015971:Gene Expression Regulation, Enzymologic; D066298:In Vitro Techniques; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D051381:Rats; D018336:Receptors, Aryl Hydrocarbon / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D018336:Receptors, Aryl Hydrocarbon