Fig. 5

HNF4α regulates HOTAIR expression in hepatocyte. a RT-qPCR analysis for the indicated epithelial and mesenchymal markers on HNF4α-silenced (for 48 h) (siHNF4α) cells, compared with control siGFP cells (siCtr). The values are calculated by the ΔCt method, expressed as fold of expression vs. the control (arbitrary value = 1) and shown as means ± S.E.M. Statistically significant differences are reported (*p < 0.05; **p < 0.01) for five independent experiments. b Western blot analysis for HNF4α on protein extracts from liver samples from three hepatocyte-specific HNF4α KO mice and three matched Cre-negative littermates. Protein amount was normalized by immunoblotting for GAPDH, as indicated. c RT-qPCR analysis for HOTAIR on liver samples from six hepatocyte-specific HNF4α KO mice and five matched Cre-negative littermates. The values are calculated by the ΔCt method, expressed as fold of expression vs. the control (arbitrary value = 1) and shown as means ± S.E.M. Statistically significant differences are reported (*p < 0.05). d RT-qPCR analysis for the indicated markers on hepatocytes treated (TGFβ) or not (NT) with TGFβ and after cytokine withdrawal (TGFβ W/D). The values are calculated by the ΔCt method, expressed as fold of expression vs. the control (arbitrary value = 1) and shown as means ± S.E.M. Statistically significant differences are reported (*p < 0.05) for three independent experiments. e qPCR analysis of ChIP assays with an anti-HNF4α antibody, or normal rabbit IgG as negative control, on chromatin from TGFβ-treated cells (+ TGFβ) or controls (− TGFβ) for 24 h and after TGFβ withdrawal, showing endogenous HNF4α binding on HOTAIR promoter consensus − 1054/− 943 (site a) and −793/−667 (site b). Timm promoter sequences were analyzed as control (Neg Ctr). Values derived from five independent experiments are reported as means ± S.E.M. and expressed as percentage of the Input chromatin (% Input). Statistically significant differences (*p < 0.05; **p < 0.01; n.s. no significant) are reported