Binding of transformed Ah receptor complex to a dioxin responsive transcriptional enhancer: evidence for two distinct heteromeric DNA-binding forms

Biochemistry. 1993 Nov 30;32(47):12841-9. doi: 10.1021/bi00210a037.


Guinea pig hepatic Ah receptor (AhR) complex was transformed in vitro to its DNA-binding form by incubation with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin). Transformed TCDD-AhR was covalently cross-linked by UV-irradiation to a bromodeoxyuridine-substituted oligonucleotide containing its specific DNA recognition site, the dioxin responsive element (DRE). Denaturing gel electrophoresis and autoradiography identified four TCDD-inducible protein-DNA complexes, with molecular masses of approximately 97, 105, and 115 kDa and a somewhat broader complex at 247 kDa. The 247-kDa complex appears to contain two distinct protein-DNA complexes of approximately 232 and 256 kDa and represents two proteins covalently cross-linked to a single DRE oligonucleotide, while the 97, 105, and 115-kDa complexes represent single protein-DRE cross-links. UV cross-linking to DRE oligonucleotides containing variable numbers of BrdU residues revealed that the 105-kDa protein, identified as the AhR ligand-binding subunit by photoaffinity labeling with a radioiodinated AhR agonist, cross-links to the DRE core consensus (5'-GCGTG-3'); the 97- and 115-kDa non-ligand-binding proteins differentially cross-link immediately 5'-ward of the core. Overall, our results not only demonstrate that the critical protein-DNA contacts which occur between the AhR complex and the DRE are made primarily by the ligand-binding subunit but also indicate that the AhR complex exists as two distinct heteromeric DNA-binding forms, containing one 105-kDa ligand-binding subunit and either one 115- or one 97-kDa non-ligand-binding subunit.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Bromodeoxyuridine / metabolism
  • Conserved Sequence
  • Cross-Linking Reagents
  • Cytosol / metabolism
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / metabolism*
  • Dioxins / chemistry
  • Enhancer Elements, Genetic* / drug effects
  • Gene Expression Regulation
  • Guinea Pigs
  • Male
  • Models, Molecular
  • Molecular Sequence Data
  • Oligodeoxyribonucleotides / chemistry
  • Oligodeoxyribonucleotides / metabolism
  • Polychlorinated Dibenzodioxins / pharmacology*
  • Protein Conformation
  • Receptors, Aryl Hydrocarbon / chemistry
  • Receptors, Aryl Hydrocarbon / metabolism*
  • Sequence Homology, Nucleic Acid
  • Ultraviolet Rays


  • Cross-Linking Reagents
  • DNA-Binding Proteins
  • Dioxins
  • Oligodeoxyribonucleotides
  • Polychlorinated Dibenzodioxins
  • Receptors, Aryl Hydrocarbon
  • 2-azido-3-iodo-7,8-dibromodibenzo-1,4-dioxin
  • Bromodeoxyuridine