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. 2017 Mar 10;7:44279. doi: 10.1038/srep44279

Figure 5. SHP represses HP-25 gene transcription by inhibiting HNF-4 binding to the promoter.

Figure 5

(a) SHP, HP-25, and albumin mRNA levels in the liver of four nonhibernating chipmunks (NHL) and four hibernating chipmunks (HL), measured by RT-qPCR. The results were normalized to the amount of mRNA of the respective genes in the NHL. The results are shown as means ± SEM. *p < 0.05. (b–g) Primary hepatocytes prepared from a nonhibernating chipmunk were transfected with a FLAG-SHP expression plasmid or an empty vector plasmid (control). In (b), whole-cell extracts from the primary hepatocytes were immunoblotted with the indicated antibodies. USF-2 was used as a loading control. In (c–e), total RNA was prepared from the primary hepatocytes (SHP or control), and the endogenous HP-25 (c), albumin (d), and HNF-4 (e) expression was assessed by RT-qPCR. The results were normalized to the amount of mRNA for each gene in control cells. Results show means ± SEM for three independent experiments. **p < 0.01. In (f and g), ChIP-qPCR was performed with chromatin from the primary hepatocytes using anti-HNF-4 (f) or anti-histone H3 (g) antibodies, and the values were normalized to the total input value. Results are shown as the fold increase over the control value. Results show means ± SEM for three independent experiments. **p < 0.01.