Gene Silencing by Chicken Ovalbumin Upstream Promoter-Transcription Factor I (COUP-TFI) Is Mediated by Transcriptional Corepressors, Nuclear Receptor-Corepressor (N-CoR) and Silencing Mediator for Retinoic Acid Receptor and Thyroid Hormone Receptor (SMRT)

Mol Endocrinol. 1997 Jun;11(6):714-24. doi: 10.1210/mend.11.6.0002.

Abstract

Chicken ovalbumin upstream promoter-transcription factors (COUP-TFs) are orphan receptors that belong to the steroid/thyroid hormone receptor (TR) superfamily and can repress the transcriptional activity of several target genes; however, the precise mechanism of this repression is unknown. Transfection of a Gal4 DNA-binding domain fused to the putative ligand-binding domain of COUP-TFI (Gal4-COUP-TFI) significantly represses the basal transcriptional activity of a reporter gene containing Gal4-binding sites. Cotransfection of COUP-TFI can relieve the Gal4-COUP-TFI repression in a dose-dependent manner. In contrast, COUP-TFI delta35, which lacks the repressor domain (the C-terminal 35 amino acids), fails to relieve this repression. This finding suggests that the repressor domain of COUP-TFI may squelch a limiting amount of corepressor in HeLa cells. In addition, increasing concentrations of TRbeta also can relieve the COUP-TFI repression in a hormone-sensitive manner. Similarly, overexpression of increasing concentration of COUP-TFI, but not COUP-TFI delta35, can squelch the silencing activity of the unliganded TRbeta. Collectively, these results indicate that COUP-TFI and TRbeta share a common corepressor(s) for their silencing activity. To determine which corepressor is involved in the COUP-TF-silencing activity, we used a yeast two-hybrid and in vitro GST pull-down assays to demonstrate that COUP-TFI can interact with the fragment of N-CoR (nuclear receptor-corepressor) encoding amino acids 921-2453 and the fragments of SMRT (silencing mediator for retinoic acid receptor and TR) encoding amino acids 29-564 and 565-1289, respectively. Interestingly, the fragment of SMRT encoding amino acids 1192-1495, which strongly interacts with TRbeta, interacts very weakly with COUP-TFI. Furthermore, overexpression of N-CoR or SMRT potentiates the silencing activity of COUP-TFI and can relieve the COUP-TFI-mediated squelching of Gal4-COUP-TFI activity. Therefore, our studies indicate that N-CoR and SMRT act as corepressors for the COUP-TFI silencing activity.

Publication types

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

MeSH terms

  • Animals
  • COUP Transcription Factor I
  • Chickens
  • DNA-Binding Proteins / genetics*
  • Gene Expression Regulation
  • HeLa Cells
  • Humans
  • Nuclear Proteins / genetics*
  • Nuclear Receptor Co-Repressor 1
  • Nuclear Receptor Co-Repressor 2
  • Promoter Regions, Genetic*
  • Receptors, Glucocorticoid / genetics*
  • Receptors, Thyroid Hormone / genetics
  • Recombinant Fusion Proteins / genetics
  • Repressor Proteins / genetics*
  • Saccharomyces cerevisiae Proteins*
  • Transcription Factors / genetics*
  • Transcription, Genetic*

Substances

  • COUP Transcription Factor I
  • DNA-Binding Proteins
  • GAL4 protein, S cerevisiae
  • NCOR1 protein, human
  • NCOR2 protein, human
  • NR2F1 protein, human
  • Nuclear Proteins
  • Nuclear Receptor Co-Repressor 1
  • Nuclear Receptor Co-Repressor 2
  • Receptors, Glucocorticoid
  • Receptors, Thyroid Hormone
  • Recombinant Fusion Proteins
  • Repressor Proteins
  • Saccharomyces cerevisiae Proteins
  • Transcription Factors