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. 2017 Mar 31;8:14866. doi: 10.1038/ncomms14866

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Copyright © 2017, The Author(s)

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(a) Colony formation assays of MCF-7 cells with Dox-inducible expression of USP9X or CEP131 in the absence or presence of Dox (upper panel). **P<0.01, Student's t-test. Colony formation assays of MCF-7 cells stably transfected with USP9X shRNA or CEP131 shRNA (lower panel). Representative images from biological triplicate experiments are shown. (b) MCF-7 cells stably transfected with USP9X shRNA or CEP131 shRNA were subjected to growth viability assay (left panel). Each bar represents the mean±s.d. for biological triplicate experiments. **P<0.01, two-way ANOVA. western blotting analysis of the indicated proteins is shown (right panel). (c) MCF-7 cells with Dox-inducible expression of CEP131 were stably transfected with USP9X shRNA for colony formation assays in the absence or presence of Dox. Representative images from biological triplicate experiments (left panel) and western blotting analysis of the indicated proteins (right panel) are shown. (d) MCF-7 infected with control lentiviruses or lentiviruses carrying shRNA against USP9X or CEP131 were transplanted on athymic mice (BALB/c; Charles River, Beijing, China) and tumour volumes were measured weekly. Each point represents the mean±s.d. for different animal measurements (n=6). **P<0.01, one-way ANOVA for tumour weight analysis and two-way ANOVA for tumour volume analysis. The levels of indicated proteins in these tumours were examined by western blotting. (e) USP9X-deficient MCF-7 tumours stably transfected with control vector or vector encoding CEP131 were transplanted onto athymic mice and tumour volumes were measured weekly. Each point represents the mean±s.d. for different animal measurements (n=6). *P<0.05; **P<0.01, one-way ANOVA for tumour weight analysis and two-way ANOVA for tumour volume analysis. The levels of indicated proteins in these tumours were examined by western blotting. (f) The xenograft tumours from four mice of each group as indicated were cultured and immunostained with antibodies against Centrin and USP9X. Scale bars, 10 μm. Representative images from two mice of each group are shown. More than one hundred cells from each xenograft were counted. Each bar represents the mean±s.d. for measurements of four xenografts in each group. *P<0.05, one-way ANOVA.