Smad Proteins Regulate Transcriptional Induction of the SM22alpha Gene by TGF-beta

Nucleic Acids Res. 2003 Feb 15;31(4):1302-10. doi: 10.1093/nar/gkg224.

Abstract

Smad proteins transduce signals from transforming growth factor-beta (TGF-beta) receptors and regulate transcription of target genes. TGF-beta is implicated in the regulation of the smooth muscle cell specific gene SM22alpha, but little is known about how Smads are involved in SM22alpha gene transcription. In this report, we demonstrate that TGF-beta activation of the SM22alpha promoter is Smad dependent in C3H10T1/2 cells, BALB 3T3 cells and neural crest Monc-1 cells. We find that the promoter region from -162 to +41 is sufficient to up-regulate the reporter gene upon TGF-beta induction. Smad3, Smad1 and Smad4 are found in TGF-beta inducible complexes that bind to a region containing a Smad binding site (SBS) and a medea box. Both the SBS and medea box are necessary for complex formation and are functionally important. Smad4 is limiting for TGF-beta induction, and Smad3, but not Smad1, significantly contributes to maximal activation. Time course luciferase assays and time course gel mobility shift assays reveal that the Smad3/4 complex is largely responsible for the immediate response of the SM22alpha promoter to TGF-beta induction, and also contributes to the maximal promoter activity. We further demonstrate that AP-1 elements contribute to induction of the SM22alpha promoter by TGF-beta.

Publication types

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

MeSH terms

  • 3T3 Cells
  • Animals
  • Base Sequence
  • Binding Sites / genetics
  • Cell Line
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Electrophoretic Mobility Shift Assay
  • Gene Expression Regulation / drug effects
  • Luciferases / genetics
  • Luciferases / metabolism
  • Mice
  • Microfilament Proteins / genetics*
  • Microfilament Proteins / metabolism
  • Molecular Sequence Data
  • Muscle Proteins / genetics*
  • Muscle Proteins / metabolism
  • Mutation
  • Nuclear Proteins / metabolism
  • Promoter Regions, Genetic / genetics
  • Protein Binding
  • Recombinant Fusion Proteins / drug effects
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Smad Proteins
  • Smad1 Protein
  • Smad3 Protein
  • Smad4 Protein
  • Trans-Activators / genetics
  • Trans-Activators / metabolism*
  • Transcription Factor AP-1 / metabolism
  • Transcription, Genetic
  • Transforming Growth Factor beta / pharmacology*

Substances

  • DNA-Binding Proteins
  • Microfilament Proteins
  • Muscle Proteins
  • Nuclear Proteins
  • Recombinant Fusion Proteins
  • Smad Proteins
  • Smad1 Protein
  • Smad1 protein, mouse
  • Smad3 Protein
  • Smad3 protein, mouse
  • Smad4 Protein
  • Smad4 protein, mouse
  • Tagln protein, mouse
  • Trans-Activators
  • Transcription Factor AP-1
  • Transforming Growth Factor beta
  • transgelin
  • Luciferases