Transcriptional activating activity of Smad4: roles of SMAD hetero-oligomerization and enhancement by an associating transactivator

Proc Natl Acad Sci U S A. 1998 Aug 18;95(17):9785-90. doi: 10.1073/pnas.95.17.9785.

Abstract

Smad4 plays a pivotal role in signal transduction of the transforming growth factor beta superfamily cytokines by mediating transcriptional activation of target genes. Hetero-oligomerization of Smad4 with the pathway-restricted SMAD proteins is essential for Smad4-mediated transcription. We provide evidence that SMAD hetero-oligomerization is directly required for the Smad4 C-terminal domain [Smad4(C)] to show its transcriptional transactivating activity; this requirement obtains even when Smad4(C) is recruited to promoters by heterologous DNA-binding domains and in the absence of the inhibitory Smad4 N-terminal domain. Defined mutations of GAL4 DNA-binding domain fusion of Smad4(C) that disrupt SMAD hetero-oligomerization suppressed transcriptional activation. Importantly, we found that an orphan transcriptional activator MSG1, a nuclear protein that has strong transactivating activity but apparently lacks DNA-binding activity, functionally interacted with Smad4 and enhanced transcription mediated by GAL4 DNA-binding domain-Smad4(C) and full-length Smad4. Transcriptional enhancement by MSG1 depended on transforming growth factor beta signaling and was suppressed by Smad4(C) mutations disrupting SMAD hetero-oligomerization or by the presence of Smad4 N-terminal domain. Furthermore, Smad4(C) did not show any detectable transactivating activity in yeast when fused to heterologous DNA-binding domains. These results demonstrate additional roles of SMAD hetero-oligomerization in Smad4-mediated transcriptional activation. They also suggest that the transcriptional-activating activity observed in the presence of Smad4 in mammalian cells may be derived, at least in part, from endogenously expressed separate transcriptional activators, such as MSG1.

MeSH terms

  • 3T3 Cells
  • Animals
  • Apoptosis Regulatory Proteins
  • Binding Sites
  • DNA-Binding Proteins*
  • Humans
  • In Vitro Techniques
  • Mice
  • Mutation
  • Nuclear Proteins / chemistry
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism
  • Protein Conformation
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins*
  • Signal Transduction
  • Smad4 Protein
  • Trans-Activators / chemistry
  • Trans-Activators / genetics
  • Trans-Activators / metabolism*
  • Transcription Factors / chemistry
  • Transcription Factors / genetics
  • Transcription Factors / metabolism
  • Transcriptional Activation
  • Transfection
  • Transforming Growth Factor beta / pharmacology

Substances

  • Apoptosis Regulatory Proteins
  • CITED1 protein, human
  • Cited1 protein, mouse
  • DNA-Binding Proteins
  • GAL4 protein, S cerevisiae
  • Nuclear Proteins
  • Recombinant Fusion Proteins
  • SMAD4 protein, human
  • Saccharomyces cerevisiae Proteins
  • Smad4 Protein
  • Smad4 protein, mouse
  • Trans-Activators
  • Transcription Factors
  • Transforming Growth Factor beta