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. 2017 Aug 28;36(50):6906–6918. doi: 10.1038/onc.2017.296

Figure 1.

Figure 1

Identification of berberine as an RXRα activator. (a) Chemical structure of berberine. (b) Berberine induced the transactivational activity of RXRα-LBD. KM12C cells were cotransfected with pBIND-RXRα-LBD and pG5luc constructs. After 24 h of transfection, cells were treated with vehicle (DMSO) or berberine (Ber) for 15 h at the indicated concentrations. The basal level of transcriptional activity in the vehicle-treated group was normalized to 1. (c) RXR antagonist UVI3003 antagonized the stimulatory effect of berberine on RXRα transactivation. The transfected cells in Figure 1b were treated with berberine (25 μm) and/or UVI3003 (10 μm) for 15 h. 9-Cis-RA (9cRA, 0.1 μm) was used as a positive control. (d) Berberine induced the transactivational activity of RXRα homodimers on RXRE in both dose- and time-dependent manners. KM12C cells were cotransfected with pGL3-RXRE and an internal control plasmid—renilla luciferase (pRL-TK) plasmid. Twenty-four hours after transfection, cells were treated with vehicle, different doses of berberine or 9-cis-RA (0.1 μm) for 15 h (left panel) or 25 μm berberine for different durations (right panel). (e) Berberine failed to induce transcriptional activity of HNF4α2. KM12C cells cotransfected with pG5luc and pBIND-HNF4α2 or pBIND-RXRα were treated with different doses of berberine for 15 h. (f) Effects of shRNA against RXRα on berberine-induced transactivational activity of RXRα homodimers and heterodimers. KM12C cells expressing control or RXRα shRNA (shCtrl or shRXRα) were cotransfected with reporter vector for RXRE, FXRE, LXRE or PPRE, together with pRL-TK plasmid. Twenty-four hours later, cells were treated with berberine for 15 h at the indicated concentration. The basal levels of transcriptional activity in the vehicle-treated groups of shCtrl and shRXRα cells were normalized to 1. All data were presented as the mean±s.e.m. of three independent experiments. Significant differences compared with vehicle controls were indicated as *P<0.05, **P<0.01 and ***P<0.001; significant differences of UVI3003 vs vehicle control (c) or shRXRα vs shCtrl control (f) at the same dose of berberine or 9-cis-RA were indicated as #P<0.05, ##P<0.01 and ###P<0.001.