Fig. 2. HIF-2α is required for the self-renewal maintenance of BCSCs by activating UPR^ER.

A The protein expression of HIF-1α and HIF-2α were measured in MCF7 and MCF7 MS cells cultured with or without PTX (3 nM) for 48 h by western blot. B The self-renewal ability was detected in HIF-2α-overexpressed (HIF-2α OE) MCF7 cells and HIF-2α-silencing (HIF-2α KD) MCF7 MS cells cultured with or without PTX (3 nM) for 48 h. Scale bar, 250 μm. C The protein expression of OCT4, NANOG, BCRP and P-gp were measured in HIF-2α OE MCF7 cells and HIF-2α KD MCF7 MS cells. D The cell viability rate was detected in HIF-2α OE MCF7 cells and HIF-2α KD MCF7 MS cells cultured with different concentrations of PTX for 48 h. Resistance index (RI) value was calculated. E The intracellular accumulation of PTX was detected in HIF-2α OE MCF7 cells and HIF-2α KD MCF7 MS cells by HPLC-MS. F The expression levels of GRP78, IRE1, XBP1s, PERK, and ATF6; phosphorylation levels of p-IRE1 and p-PERK were measured in MCF7 cells, MCF7 MS cells, HIF-2α OE MCF7 cells and HIF-2α KD MCF7 MS cells. G The self-renewal ability rescued in GRP78-overexpressing and HIF-2α-silencing (GRP78 OE + HIF-2α KD) MCF7 MS cells, compared with HIF-2α KD MCF7 MS cells. Scale bar, 250 μm. H The expression levels of GRP78, OCT4 and P-gp were measured in GRP78 OE + HIF-2α KD MCF7 MS cells. I The intracellular accumulation of PTX was detected in GRP78 OE + HIF-2α KD MCF7 MS cells by HPLC-MS. J The cells viability rate was detected in GRP78 OE + HIF-2α KD MCF7 MS cells cultured with different concentrations of PTX (left panel), the related IC₅₀ of PTX were calculated (right panel). NS, non-significant; #P < 0.05, ##P < 0.01, ###P < 0.001, compared to treatment without PTX, *P < 0.05, **P < 0.01, ***P < 0.001, compared to Ctrl/shCtrl/shHIF-2α; Student’s t test, two-way ANOVA test. Error bars, mean ± SD (n = 3).