Characterization of the molecular and structural properties of the transformed and nuclear aryl hydrocarbon (Ah) receptor complexes by proteolytic digestion

M. Santostefano, S. Safe

Research output: Contribution to journalArticle

7 Scopus citations

Abstract

Ligand-dependent differences in the molecular properties of the transformed cytosolic and nuclear aryl hydrocarbon receptor (AhR) were investigated using the proteolytic clipping band shift assay. AhR complexes were incubated with [32P]dioxin responsive element (DRE) (26-mer) or bromodeoxyuridine (BrdU)-DRE and the resulting protein-DNA or crosslinked protein-DNA complexes were treated with trypsin or V8 protease and analyzed by electrophoresis. The results showed that for several different AhR ligands including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 2,3,7,8- tetrachlorodibenzofuran, 1,2,7,8-tetrachlorodibenzofuran and α- naphthoflavone, the pattern of degraded protein-DNA products were similar using transformed cytosolic or nuclear AhR complexes. In contrast, the proteolytic clipping band shift assay showed that there were significant differences in the pattern of degraded protein-DNA products using nuclear AhR complexes derived from mouse Hepa 1c1c7 cells treated with TCDD or 6-methyl- 1,3,8-trichlorodibenzofuran (MCDF). The differences detected in this in vitro assay parallel the in vivo and in vitro activities of these compounds in which TCDD is a potent AhR agonist whereas MCDF is a partial AhR agonist and antagonist.

Original languageEnglish (US)
Pages (from-to)221-240
Number of pages20
JournalChemico-Biological Interactions
Volume100
Issue number3
DOIs
StatePublished - May 6 1996

Keywords

  • Aryl hydrocarbon receptor
  • Ligand
  • Proteolytic digestion
  • Trypsin
  • V8 protease

ASJC Scopus subject areas

  • Toxicology

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