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Related Experiment Videos

Smad8/9 Is Regulated Through the BMP Pathway.

Yuko Katakawa1, Masayuki Funaba2, Masaru Murakami1

  • 1Laboratory of Molecular Biology, Azabu University School of Veterinary Medicine, Sagamihara, 252-5201, Japan.

Journal of Cellular Biochemistry
|January 11, 2016
PubMed
Summary

Bone morphogenetic protein 4 (BMP4) directly up-regulates Smad8/9 gene expression via the BMP type I receptor pathway. This regulation occurs transcriptionally, involving specific BMP-responsive elements bound by phosphorylated Smad proteins.

Keywords:
BMPSMAD8/9TRANSCRIPTION

Related Experiment Videos

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Signal Transduction

Background:

  • Transforming growth factor-β (TGF-β) family signaling involves Smad-dependent and -independent pathways.
  • Receptor-regulated Smad (R-Smad) activity is modulated by phosphorylation, dephosphorylation, and gene induction of inhibitory Smads (I-Smads).
  • Regulation of R-Smad gene expression remains largely uncharacterized.

Purpose of the Study:

  • To investigate the regulation of Smad8/9 gene expression.
  • To elucidate the role of Bone Morphogenetic Protein 4 (BMP4) in Smad8/9 expression.
  • To identify the signaling pathway and regulatory elements involved in BMP4-induced Smad8/9 expression.

Main Methods:

  • Treatment of various cell lines (C2C12, H9c2, 3T3-L1, HepG2, B16, primary fibroblasts) with BMP4.
  • Assessment of Smad8/9 expression levels.
  • Use of cycloheximide to inhibit protein synthesis.
  • Use of LDN-193189, a BMP type I receptor inhibitor.
  • Reporter assays to identify BMP-responsive elements (BREs).
  • Chromatin immunoprecipitation to detect Smad protein binding to DNA.

Main Results:

  • BMP4 treatment significantly increased Smad8/9 mRNA expression across multiple cell types.
  • BMP4-induced Smad8/9 upregulation was insensitive to cycloheximide but sensitive to LDN-193189, indicating a direct transcriptional mechanism via BMP type I receptors.
  • A specific region (nt -121 to -44) within the Smad8/9 gene promoter contained BMP-responsive elements (BREs).
  • Phosphorylated Smad1/5/8/9 proteins directly bound to these BREs.

Conclusions:

  • Smad8/9 is a unique R-Smad whose gene expression is directly regulated by the BMP signaling pathway at the mRNA level.
  • BMP4 transcriptionally activates the Smad8/9 gene through specific BREs.
  • This study reveals a novel regulatory mechanism for R-Smad expression within the BMP signaling cascade.