Fig. 6: ZNF32 and GPER regulate breast cancer stem cell populations by activating the ERK pathway.

(a) Western analysis was used to detect ERK activation. Quantification of ZNF32 and pERK expression in 3 independent experiments. (b) Western analysis was used to detect ERK activation in ZNF32 over-expressing cells with or without GPER interference. Quantification of ZNF32 and pERK expression levels from 3 independent experiments. (c) ERK inhibitor U0126 (20 μM) or DMSO (control) was added to ZR-75-30 cells over-expressing ZNF32, followed by incubation for 24 h. Western analysis was used to detect ERK activation. Quantification of ZNF32 and pERK levels from 3 independent experiments. (d) qPCR analysis was used to measured ALDH1 gene expression in ZNF32 over-expressing or vector control ZR-75-30 cells treated with or without U0126 or DMSO for 24 h. (e) qPCR analysis was used to detect OCT4, Nanog and KLF4 gene expression in ZNF32 over-expressing or vector control ZR-75-30 cells treated with or without U0126 or DMSO for 24 h. (f) (g) ZR-75-30 cells with ZNF32 over-expression or knockdown were plated in 96-well plates. After 24 h of incubation, pretreatment with 20 μM U0126 was performed for 2 h before the addition of 10 ng/ml or 20 ng/ml Taxol or DMSO (control), and MTT analysis was used to detect cell viability