Transcriptional regulation of SM22α by Wnt3a: Convergence with TGFβ1/Smad signaling at a novel regulatory element

Shawn L. Shafer, Dwight A. Towler

Research output: Contribution to journalArticle

32 Scopus citations

Abstract

The role of canonical Wnt signaling in myofibroblast biology has not been fully investigated. The C3H10T1/2 mesenchymal cell line recapitulates myofibroblast differentiation in vitro and in vivo, including SM22α expression. Using this model, we find that Wnt3a upregulates SM22α in concert with TGFβ1. Wnt1, Wnt5a and BMP2 could not replace Wnt3a and TGFβ1 signals. Chromatin immunoprecipitation identified that Wnt3a enhances both genomic SM22α histone H3 acetylation and β-catenin association, hallmarks of transcriptional activation. By analyzing a series of SM22α promoter-luciferase (LUC) reporter constructs, we mapped Wnt3a-regulated DNA transcriptional activation to nucleotides - 213 to - 192 relative to the transcription initiation site. In gel shift assays, DNA-protein complexes assembled on this element were disrupted with antibodies to β-catenin, Smad2/3, and TCF7, confirming the participation of known Wnt3a and TGFβ transcriptional mediators. Mutation of a CAGAG motif within this region abrogated recognition by these DNA binding proteins. Wnt3a treatment increased Smad2/3 binding to this element. Mutation of the cognate within the context of the native 0.44 kb SM22α promoter resulted in a 70% decrease in transcription, and reduced Wnt3a + TGFβ1 induction. A concatamer of SM22α [- 213 to - 192] conveyed Wnt3a + TGFβ1 activation to the unresponsive RSV promoter. Dominant negative TCF inhibited SM22α [- 213 to - 192]x6 RSVLUC activation. Moreover, ICAT (inhibitor of β-catenin and TCF) decreased while TCF7L2 and β-catenin enhanced 0.44 kb SM22α promoter induction by Wnt3a + TGFβ1. RNAi "knockdown" of β-catenin inhibited Wnt3a induction of SM22α. Thus, Wnt/β-catenin signaling interacts with TGFβ/Smad pathways to control SM22α gene transcription.

Original languageEnglish (US)
Pages (from-to)621-635
Number of pages15
JournalJournal of Molecular and Cellular Cardiology
Volume46
Issue number5
DOIs
StatePublished - May 1 2009

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Keywords

  • Msx2
  • Myofibroblast
  • Type II diabetes
  • Vascular calcification
  • Wnt
  • β-catenin

ASJC Scopus subject areas

  • Molecular Biology
  • Cardiology and Cardiovascular Medicine

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