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. Author manuscript; available in PMC: 2018 Apr 24.
Published in final edited form as: Biochem J. 2017 Apr 24;474(9):1531–1546. doi: 10.1042/BCJ20160943

Figure 6. A TGF-β-activated transcriptional complex induces PPARG transcriptional repression by recruiting HDAC1.

Figure 6

IMR-90 human lung fibroblasts were treated with or without TGF-β (2 ng/ml). Following treatment, nuclear extracts were obtained and were incubated with the indicated PPARG promoter or SMAD reporter promoter biotinylated plasmid-coupled beads. Bead-bound biotinylated plasmid-protein complexes were eluted and (A) subjected to Western blot to identify the presence of p300 and HDAC1. (B) A histone deacetylation activity assay was performed with eluted samples and recombinant HDAC1 in the presence and absence of TSA using 2 μg of acetylated (K16) histone H4 in HDAC assay buffer and were then subjected to Western blot to identify the amount of deacetylation under indicated conditions. The results were reproduced in two to three independent experiments. (C) Schematic showing how TGF-β-activated R-SMADs can either activate target gene expression by recruitment of coactivators and other transcription factors that have histone acetylation activity or repress the target gene by recruitment of transcriptional corepressors and transcription factors that have histone deacetylation activity.