title: TGFβ1 enhances MAD1 expression and stimulates promoter-bound Pol II phosphorylation: basic functions of C/EBP, SP and SMAD3 transcription factors.
abstract: BACKGROUND
  The MAD1 protein, a member of the MYC/MAX/MAD network of transcriptional regulators, controls cell proliferation, differentiation and apoptosis. MAD1 functions as a transcriptional repressor, one direct target gene being the tumor suppressor PTEN. Repression of this gene is critical to mediate the anti-apoptotic function of MAD1. Under certain conditions it also antagonizes the functions of the oncoprotein MYC. Previous studies have demonstrated that MAD1 expression is controlled by different cytokines and growth factors. Moreover we have recently demonstrated that the MAD1 promoter is controlled by the cytokine granulocyte colony-stimulating factor (G-CSF) through the activation of STAT3, MAP kinases and C/EBP transcription factors.
  
  
  RESULTS
  We observed that in addition to G-CSF, the cytokine transforming growth factor β (TGFβ1) rapidly induced the expression of MAD1 mRNA and protein in promyelocytic tumor cells. Moreover we found that C/EBP and SP transcription factors cooperated in regulating the expression of MAD1. This cooperativity was dependent on the respective binding sites in the proximal promoter, with the CCAAT boxes being bound by C/EBPα/β heterodimers. Both C/EBP and SP transcription factors bound constitutively to DNA without obvious changes in response to TGFβ1. In addition SMAD3 stimulated the MAD1 reporter, cooperated with C/EBPα and was bound to the core promoter region. Thus SMAD3 appears to be a potential link between TGFβ1 signaling and C/EBP regulated promoter activity. Moreover TGFβ1 stimulated the phosphorylation of polymerase II at serine 2 and its progression into the gene body, consistent with enhanced processivity.
  
  
  CONCLUSIONS
  Our findings suggest that C/EBP and SP factors provide a platform of transcription factors near the core promoter of the MAD1 gene that participate in mediating signal transduction events emanating from different cytokine receptors. SMAD3, a target of TGFβ1 signaling, appears to be functionally relevant. We suggest that a key event induced by TGFβ1 at the MAD1 promoter is the recruitment or activation of cofactors, possibly in complex with C/EBP, SP, and SMAD3 transcriptional regulators, that control polymerase activity.
authors.affiliation: Institute of Biochemistry and Molecular Biology, Medical School, RWTH Aachen University, 52057 Aachen, Germany.
authors.forename: Nadine
authors.initials: N
authors.lastname: Hein
authors.forename: Kan
authors.initials: K
authors.lastname: Jiang
authors.forename: Christian
authors.initials: C
authors.lastname: Cornelissen
authors.forename: Bernhard
authors.initials: B
authors.lastname: Lüscher
mesh_terms: D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000235:genetics / Q000378:metabolism*; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D004257:DNA Polymerase II / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D051900:Smad3 Protein / Q000235:genetics / Q000378:metabolism*; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation
chemical_list: D022763:CCAAT-Enhancer-Binding Protein-alpha; D022782:CCAAT-Enhancer-Binding Protein-beta; D018797:Cell Cycle Proteins; C501384:MAD1L1 protein, human; D009687:Nuclear Proteins; D051900:Smad3 Protein; D016329:Sp1 Transcription Factor; C508960:TGFB1 protein, human; D053773:Transforming Growth Factor beta1; D004257:DNA Polymerase II