Smad2 and PEA3 cooperatively regulate transcription of response gene to complement 32 in TGF-β-induced smooth muscle cell differentiation of neural crest cells.
Response gene to complement 32 (RGC-32) is activated by transforming growth factor- β (TGF-β) and plays an important role in smooth muscle cell (SMC) differentiation from neural crest Monc-1 cells. The molecular mechanism governing TGF-β activation of RGC-32, however, remains to be determined. The present studies indicate that TGF-β regulates ... RGC-32 gene transcription. Sequence analysis revealed a Smad binding element (SBE) located in the region from -1344 to -1337 bp upstream of the transcription start site of RGC-32 gene. A polyomavirus enhancer activator (PEA3) binding site is adjacent to the SBE. Mutation at either SBE or PEA3 site significantly inhibited RGC-32 promoter activity. Mutations at both sites completely abolished TGF-β-induced promoter activity. Biochemically, TGF-β stimulated recruitment of Smad2, Smad4, and PEA3 to the RGC-32 promoter, as revealed by gel shift and chromatin immunoprecipitation analyses. Functionally, Smad2, but not Smad3, activated RGC-32 promoter. PEA3 appeared to enhance Smad2 activity. In agreement with their function, Smad2, but not Smad3, physically interacted with PEA3. In TGF-β-induced SMC differentiation of Monc-1 cells, knockdown of Smad2 by short hairpin RNA resulted in downregulation of RGC-32 and SMC marker genes. The downregulation of SMC markers, however, was rescued by exogenously introduced RGC-32. These results demonstrate that Smad2 regulation of RGC-32 transcription is essential for SMC differentiation from neural crest cells.
Mesh Terms:
Animals, Binding Sites, Biological Markers, Cell Differentiation, Cell Line, Chromatin Immunoprecipitation, Electrophoretic Mobility Shift Assay, Embryonic Stem Cells, Gene Expression Regulation, Developmental, Mice, Mutation, Myocytes, Smooth Muscle, Neural Crest, Nuclear Proteins, Promoter Regions, Genetic, RNA Interference, Signal Transduction, Smad2 Protein, Smad3 Protein, Transcription Factors, Transcriptional Activation, Transfection, Transforming Growth Factor beta1
Animals, Binding Sites, Biological Markers, Cell Differentiation, Cell Line, Chromatin Immunoprecipitation, Electrophoretic Mobility Shift Assay, Embryonic Stem Cells, Gene Expression Regulation, Developmental, Mice, Mutation, Myocytes, Smooth Muscle, Neural Crest, Nuclear Proteins, Promoter Regions, Genetic, RNA Interference, Signal Transduction, Smad2 Protein, Smad3 Protein, Transcription Factors, Transcriptional Activation, Transfection, Transforming Growth Factor beta1
Am. J. Physiol., Cell Physiol.
Date: Aug. 01, 2011
PubMed ID: 21613609
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