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TGFβ1 enhances MAD1 expression and stimulates promoter-bound Pol II phosphorylation: basic functions of C/EBP, SP and

Nadine Hein1, Kan Jiang, Christian Cornelissen

  • 1Institute of Biochemistry and Molecular Biology, Medical School, RWTH Aachen University, 52057 Aachen, Germany.

BMC Molecular Biology
|February 25, 2011
PubMed
Abstract

Insights

Transforming growth factor beta 1 (TGFβ1) rapidly induces MAD1 expression by engaging C/EBP and SP transcription factors, with SMAD3 acting as a key mediator. This cytokine signaling pathway regulates cell proliferation and apoptosis through the MAD1 gene.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Gene Regulation

Background:

  • MAD1 protein, a transcriptional repressor in the MYC/MAX/MAD network, regulates cell proliferation, differentiation, and apoptosis.
  • MAD1 represses the tumor suppressor PTEN, mediating its anti-apoptotic function and antagonizing the oncoprotein MYC.
  • MAD1 expression is controlled by cytokines like granulocyte colony-stimulating factor (G-CSF) via STAT3, MAP kinases, and C/EBP transcription factors.

Purpose of the Study:

  • To investigate the role of transforming growth factor beta 1 (TGFβ1) in regulating MAD1 expression.
  • To identify transcription factors involved in TGFβ1-mediated induction of MAD1.
  • To elucidate the mechanism by which TGFβ1 signaling influences MAD1 gene transcription.

Main Methods:

  • Analysis of MAD1 mRNA and protein levels following TGFβ1 treatment in promyelocytic tumor cells.
  • Reporter gene assays to assess promoter activity.
  • Electrophoretic mobility shift assays (EMSAs) to study transcription factor binding to the MAD1 promoter.
  • Co-immunoprecipitation to investigate protein-protein interactions.
  • Analysis of RNA polymerase II phosphorylation and progression.

Main Results:

  • TGFβ1 rapidly induced MAD1 mRNA and protein expression.
  • C/EBP and SP transcription factors cooperated to regulate MAD1 expression, binding to CCAAT boxes and the core promoter region, respectively.
  • SMAD3, a TGFβ1 signaling target, stimulated MAD1 reporter activity, cooperated with C/EBPα, and bound to the core promoter.
  • TGFβ1 enhanced RNA polymerase II phosphorylation and gene body progression.

Conclusions:

  • C/EBP and SP factors form a platform at the MAD1 core promoter for cytokine receptor signaling.
  • SMAD3 is a key mediator linking TGFβ1 signaling to C/EBP-regulated MAD1 promoter activity.
  • TGFβ1 induces MAD1 expression by recruiting cofactors that modulate polymerase activity, involving C/EBP, SP, and SMAD3.

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