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

Endocrinology. 2002 Apr;143(4):1280-90. doi: 10.1210/endo.143.4.8748.

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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adrenal Cortex / enzymology*
  • Amino Acid Sequence
  • Blotting, Western
  • Cell Nucleus / genetics
  • Cells, Cultured
  • Cycloheximide / pharmacology
  • DNA-Binding Proteins / physiology*
  • Fushi Tarazu Transcription Factors
  • Gene Expression Regulation, Enzymologic / genetics
  • Histone Deacetylases / genetics
  • Homeodomain Proteins
  • Humans
  • Immunoenzyme Techniques
  • Molecular Sequence Data
  • Nuclear Proteins / genetics*
  • Plasmids / genetics
  • Promoter Regions, Genetic / genetics
  • Protein Synthesis Inhibitors / pharmacology
  • Receptors, Cytoplasmic and Nuclear
  • Serine-Arginine Splicing Factors
  • Steroid 17-alpha-Hydroxylase / genetics*
  • Steroid 17-alpha-Hydroxylase / metabolism*
  • Steroidogenic Factor 1
  • Transcription Factors / physiology*
  • Transcription, Genetic / genetics
  • Transcriptional Activation / genetics*
  • Transcriptional Activation / physiology*

Substances

  • DNA-Binding Proteins
  • Fushi Tarazu Transcription Factors
  • Homeodomain Proteins
  • Nuclear Proteins
  • Protein Synthesis Inhibitors
  • Receptors, Cytoplasmic and Nuclear
  • Sfrs11 protein
  • Steroidogenic Factor 1
  • Transcription Factors
  • Serine-Arginine Splicing Factors
  • Cycloheximide
  • Steroid 17-alpha-Hydroxylase
  • Histone Deacetylases