Fig. 2: TGF-β-induced downregulation of C/EBPα is mediated by the transcription factor Smad3. | Nature Communications

Fig. 2: TGF-β-induced downregulation of C/EBPα is mediated by the transcription factor Smad3.

From: C/EBPɑ is crucial determinant of epithelial maintenance by preventing epithelial-to-mesenchymal transition

Fig. 2

a Visualization of SMAD3 CHIP-seq profile within the genomic region surrounding the CEBPA locus. Stars indicate enriched picks upon 16 h of TGF-β stimulation. Part of the SMAD3-binding regions sequence is displayed, showing in bold SMAD3 known primary binding motif (C/TC/TA/CGACA/G). b Chromatin immunoprecipitation was performed in TGF-β treated or untreated HMLE cells using Smad3 antibody and according to “Methods”. qRT-PCR was performed using specific primers targeting enriched areas (regions 1 and 2) on CEBPA locus. Data represented as mean ± SD of three independent experiments, normalized for a negative region. p-values were calculated using unpaired one-tailed Student’s t test. *p < 0.05. c The results from qRT-PCR showing SMAD3 mRNA levels in HMLE cells expressing shRNA control (SCR) or two independent shRNAs targeting human SMAD3 (shSMAD3_1 and shSMAD3_2). Data represented as mean ± SD of three independent experiments. p-values were calculated using unpaired two-tailed Student’s t test. ****p < 0.0001. d HMLE cells expressing shSCR or shSMAD3_2 were left untreated or treated with TGF-β for 4 or 8 h. qRT-PCR analysis displaying the effect of TGF-β stimulation on CEBPA mRNA levels in the presence of SMAD3 knockdown. Data represented as mean ± SD of three independent experiments. p-values were calculated using two-way ANOVA with Tukey’s multiple comparisons test. neg negatived, h hours, FC fold change, β2m beta-2-microglobulin, UNT untreated.

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