title: Transcriptional activation of human CYP17 in H295R adrenocortical cells depends on complex formation among p54(nrb)/NonO, protein-associated splicing factor, and SF-1, a complex that also participates in repression of transcription.
abstract: The first 57 bp upstream of the transcription initiation site of the human CYP17 (hCYP17) gene are essential for both basal and cAMP-dependent transcription. EMSA carried out by incubating H295R adrenocortical cell nuclear extracts with radiolabeled -57/-38 probe from the hCYP17 promoter showed the formation of three DNA-protein complexes. The fastest complex contained steroidogenic factor (SF-1) and p54(nrb)/NonO, the intermediate complex contained p54(nrb)/NonO and polypyrimidine tract-binding protein-associated splicing factor (PSF), and the slowest complex contained an SF-1/PSF/p54(nrb)/NonO complex. (Bu)(2)cAMP treatment resulted in a cAMP-inducible increase in the binding intensity of only the upper complex and also activated hCYP17 gene transcription. SF-1 coimmunoprecipitated with p54(nrb)/NonO, indicating direct interaction between these proteins. Functional assays revealed that PSF represses basal transcription. Further, the repression of hCYP17 promoter-reporter construct luciferase activity resulted from PSF interacting with the corepressor mSin3A. Trichostatin A attenuated the inhibition of basal transcription, suggesting that a histone deacetylase interacts with the SF-1/PSF/p54(nrb)/NonO/mSin3A complex. Our studies lend support to the idea that the balance between transcriptional activation and repression is essential in the control of adrenocortical steroid hormone biosynthesis.
authors.affiliation: Department of Biochemistry and Center in Toxicology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0146, USA. marion@toxicology.mc.vanderbilt.edu
authors.forename: Marion B
authors.initials: MB
authors.lastname: Sewer
authors.forename: Viet Q
authors.initials: VQ
authors.lastname: Nguyen
authors.forename: Ching-Jung
authors.initials: CJ
authors.lastname: Huang
authors.forename: Philip W
authors.initials: PW
authors.lastname: Tucker
authors.forename: Norio
authors.initials: N
authors.lastname: Kagawa
authors.forename: Michael R
authors.initials: MR
authors.lastname: Waterman
mesh_terms: D000302:Adrenal Cortex / Q000201:enzymology*; D000595:Amino Acid Sequence; D015153:Blotting, Western; D002467:Cell Nucleus / Q000235:genetics; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000502:physiology*; D051717:Fushi Tarazu Transcription Factors; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D006655:Histone Deacetylases / Q000235:genetics; D018398:Homeodomain Proteins; D006801:Humans; D007124:Immunoenzyme Techniques; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D018160:Receptors, Cytoplasmic and Nuclear; D000068103:Serine-Arginine Splicing Factors; D013254:Steroid 17-alpha-Hydroxylase / Q000235:genetics* / Q000378:metabolism*; D054339:Steroidogenic Factor 1; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic / Q000235:genetics; D015533:Transcriptional Activation / Q000235:genetics* / Q000502:physiology*
chemical_list: D004268:DNA-Binding Proteins; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; D011500:Protein Synthesis Inhibitors; D018160:Receptors, Cytoplasmic and Nuclear; C101745:Sfrs11 protein; D054339:Steroidogenic Factor 1; D014157:Transcription Factors; D000068103:Serine-Arginine Splicing Factors; D003513:Cycloheximide; D013254:Steroid 17-alpha-Hydroxylase; D006655:Histone Deacetylases