Objective To investigate the effects of B cell lymphoma factor-3(Bcl-3)on estrogen receptor α(ERα)signaling pathway and cell proliferation in breast cancer.Methods MCF-7 cells at the logarithmic growth stage were randomly divided into the siControl group and the siBcl-3#1 group.The cells were added with ControlsiRNA and siBcl-3#1siRNA,respectively,and transfected with LipofectamineTM 2000 for 5 hours.MCF-7 cells in the siControl group and siBcl-3#1 group were obtained after stable transfection for 48 hours.The expressions of Bcl-3,growth regulation by estrogen in breast cancer 1(GREB1),PDZ domain protein 1(PDZK1),and Erα target gene trefoil factor 1(TFF1)mRNA in MCF-7 cells were detected by quantitative real-time polymerase chain reaction.The expressions of Bcl-3 and ERα protein in MCF-7 cells were detected by Western blot assay,and the proliferation of MCF-7 cells was detected by cell counting kit-8.HEK293T cells were randomly divided into the Input group(positive control group),IP group(co-immunoprecipitation group),and IgG group(negative control group).After co-transfected with Bcl-3 and ERα plasmids for 48 hours,the cells were lysed with 100 μL co-immunoprecipitation(Co-IP)cell lysate on ice,and the supernatant was separated.In the Input group,90 µL supernatant was taken and added with 30 μL 4 × Loading buffer.In the IP group,450μL supernatant was taken and added with 40 μL protein A/G and 0.5 μL green fluorescent protein antibody.In the IgG group,450 μL supernatant was taken and added with 0.5 μL IgG and 30 μL protein A/G magnetic agarose beads.The binding effect of Bcl-3 and ERα was detected by Co-IP assay.MCF-7 cells at the logarithmic growth stage were selected,and the co-localization of Bcl-3 and ERα was detected by an immunofluorescence co-localization experiment.Results The relative expression levels of Bcl-3,GREB1,TFF1 and PDZK1 mRNA in MCF-7 cells in the si-Bcl-3#1 group were significantly lower than those in the siControl group(P＜0.05).The relative expression levels of Bcl-3 and ERα protein in MCF-7 cells in the si-Bcl-3#1 group were significantly lower than those in the siControl group(P＜0.05).There was no significant difference in the proliferation ability of MCF-7 cells between the si-Bcl-3#1 group and the siControl group at 0 h and 24 h after culture(P＞0.05).The proliferation ability of MCF-7 cells in the si-Bcl-3#1 group was significantly lower than that in the siControl group at 48 h and 72 h after culture(P＜0.05).The results of the Co-IP assay showed that there was interaction between Bcl-3 and ERα.The results of the immunofluorescence co-localization experiment showed that the co-localization of Bcl-3 and ERα existed.Conclusion Bcl-3 is highly expressed in breast cancer cells,which can enhance the conduction activity of the ERα signaling pathway and promote the proliferation of ERα positive breast cancer cells.
B cell lymphoma factor-3breast cancerestrogen receptor αcell proliferation
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