Background： MicroRNA-206 （miR-206） and connexin 43 （Cx43） are related with the distant metastasis of breast cancer.It remains unclear whether the regulatory effect of miR-206 on Cx43 is involved in metastasis of ...Background： MicroRNA-206 （miR-206） and connexin 43 （Cx43） are related with the distant metastasis of breast cancer.It remains unclear whether the regulatory effect of miR-206 on Cx43 is involved in metastasis of breast cancer.Methods： Using quantitative real-time polymerase chain reaction and Western blot, the expressions of miR-206 and Cx43 were determined in breast cancer tissues, hepatic and pulmonary metastasis （PM）, and cell lines （MCF-10A, MCF-7, and MDA-MB-231）.MCF-7/MDA-M-231 cells were transfected with lentivirus-shRNA vectors to enhance/inhibit miR-206, and then Cx43 expression was observed.Cell counting kit-8 assay and Transwell method were used to detect their changes in proliferation, migration, and invasion activity.The mutant plasmids of Cx43-3＆#39; untranslated region （3＆#39;UTR） at position 478-484 and position 1609-1615 were constructed.Luciferase reporter assay was performed to observe the effects of miR-206 on luciferase expression of different mutant plasmids and to confirm the potential binding sites of Cx43.Results： Cx43 protein expression in hepatic and PM was significantly higher than that in the primary tumor, while no significant difference was showed in messenger RNA （mRNA） expression.MiR-206 mRNA expression in hepatic and PM was significantly lower than that in the primary tumor.Cx43 mRNA and protein levels, as well as cell proliferation, migration, and invasion capabilities, were all significantly improved in MDA-MB-231 cells after reducing miR-206 expression but decreased in MCF-7 cells after elevating miR-206 expression, which demonstrated a significantly negative correlation between miR-206 and Cx43 expression （P =0.03）.MiR-206 can drastically decrease Cx43 expression of MCF-7 cells but exerts no effects on Cx43 expression in 293 cells transfected with the Cx43 coding region but the lack of Cx43-3＆#39;UTR, suggesting that Cx43-3＆#39;UTR may be the key in Cx43 regulated by miR-206.Luciferase expression showed that the inhibition efficiency was reduced by 46.80% in position 478-484 mutant, 16.72% in position 1609-1615 mutant;the inhibition was totally disappeared in double mutant （P =0.02）.Conclusions： MiR-206 can regulate the expression of Cx43, the cytobiological activity, and the metastasis of breast cancer through binding to the two binding sites in Cx43-3＆#39;UTR： position 478-484 and position 1609-1615.展开更多
文摘Background： MicroRNA-206 （miR-206） and connexin 43 （Cx43） are related with the distant metastasis of breast cancer.It remains unclear whether the regulatory effect of miR-206 on Cx43 is involved in metastasis of breast cancer.Methods： Using quantitative real-time polymerase chain reaction and Western blot, the expressions of miR-206 and Cx43 were determined in breast cancer tissues, hepatic and pulmonary metastasis （PM）, and cell lines （MCF-10A, MCF-7, and MDA-MB-231）.MCF-7/MDA-M-231 cells were transfected with lentivirus-shRNA vectors to enhance/inhibit miR-206, and then Cx43 expression was observed.Cell counting kit-8 assay and Transwell method were used to detect their changes in proliferation, migration, and invasion activity.The mutant plasmids of Cx43-3＆#39; untranslated region （3＆#39;UTR） at position 478-484 and position 1609-1615 were constructed.Luciferase reporter assay was performed to observe the effects of miR-206 on luciferase expression of different mutant plasmids and to confirm the potential binding sites of Cx43.Results： Cx43 protein expression in hepatic and PM was significantly higher than that in the primary tumor, while no significant difference was showed in messenger RNA （mRNA） expression.MiR-206 mRNA expression in hepatic and PM was significantly lower than that in the primary tumor.Cx43 mRNA and protein levels, as well as cell proliferation, migration, and invasion capabilities, were all significantly improved in MDA-MB-231 cells after reducing miR-206 expression but decreased in MCF-7 cells after elevating miR-206 expression, which demonstrated a significantly negative correlation between miR-206 and Cx43 expression （P =0.03）.MiR-206 can drastically decrease Cx43 expression of MCF-7 cells but exerts no effects on Cx43 expression in 293 cells transfected with the Cx43 coding region but the lack of Cx43-3＆#39;UTR, suggesting that Cx43-3＆#39;UTR may be the key in Cx43 regulated by miR-206.Luciferase expression showed that the inhibition efficiency was reduced by 46.80% in position 478-484 mutant, 16.72% in position 1609-1615 mutant;the inhibition was totally disappeared in double mutant （P =0.02）.Conclusions： MiR-206 can regulate the expression of Cx43, the cytobiological activity, and the metastasis of breast cancer through binding to the two binding sites in Cx43-3＆#39;UTR： position 478-484 and position 1609-1615.
