Journal: Nature Communications
Article Title: EZH2-H3K27me3 mediated KRT14 upregulation promotes TNBC peritoneal metastasis
doi: 10.1038/s41467-022-35059-x
Figure Lengend Snippet: A Immunoblot analysis for EZH2, H3K27me3, β-actin in control, and EZH2 OE HCC1806 cells. B Control and EZH2 Y641-F cells were analyzed for expression of EZH2, H3K27me3, KRT14, and β-actin by Immunoblot. C Immunoblot analysis for EZH2, H3K27me3, β-actin in control, and EZH2 EZH1SET HCC1806 cells. D The control and EZH2 Y641-F cells were treated with a 10 μM dose of EPZ6438 and performed qPCR analysis for KRT14 expression. Data points are the mean of triplicate readings of samples; error bars, ±S.D, compared to control * P = 0.0001 and Y641-F # P = 0.0001, respectively, one-way ANOVA, Turkey’s multiple comparisons test. E The control and EZH2 Y641-F cells were treated with a 10 μM dose of EPZ6438 and analyzed for expression of EZH2, H3K27me3, KRT14, and β-actin by immunoblot. F The vehicle control and Tet-ON EZH2-YFP OE HCC1806 cells were analyzed for expression of EZH2 WT, EZH2-YFP, H3K27me3, KRT14, and β-actin by immunoblot. G The HCC1806 cells were treated either for vehicle control or 10 μM dose of EPZ6438 for the analysis of the expression of KRT14 by RT-qPCR. Data points are the mean of triplicate readings of samples; error bars, ±S.D, * P = 0.0001 compared to control cells, student’s t -test (two-sided). H The HCC1806 and MDAMB468 cells were either treated for a vehicle or 10 μM EPZ6438 to check the expression of EZH2, H3K27me3, KRT14, and β-actin by immunoblot. I HCC-1806 cells were treated with either vehicle or 10 μM EPZ6438 treatments for 24 h, co-stained with H3K27me3 (red) and KRT14 (green) antibodies, and analyzed under a confocal microscope. Scale bar, 50 μm. J , K 4T-1 cells were isolated from primary tumors and metastatic spleen and subjected to confocal analysis after having either co-staining ( J ) with H3K27me3 (red) and KRT14 (green) or single ( K ) staining with Ki-67 (red) antibodies. In both cases, DAPI was used to stain the nuclei. Scale bar 20 and 5 μm. Source data are provided as a Source Data file.
Article Snippet: MSCV (cat# 24828), MSCV EZH2 ΔSET-Hygro (cat# 49403) EZH2-Y641-F (cat# 80077), pTRIPZ M)-YFP-EZH2 (cat# 82511), MSCV-EZH2OE (cat#75125), MSCV–EZH2 EZH1SET (cat#75126) were procured from Addgene USA.
Techniques: Western Blot, Control, Expressing, Quantitative RT-PCR, Staining, Microscopy, Isolation
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