Figure 7A PC3-LN4 Naïve and CRISPR/Cas9 genome-edited EDC3 S161A (SA), S161D (Fredriksson et al.), and EDC3 knock-out (KO) cell lines were plated (4000 cells/well in 24 well plates) and imaged every 12 h using phase contrast in the IncuCyte ZOOM platform. The graph shown was generated with IncuCyte Basic Software following 5 days of growth. The experiments were done in triplicates. Values are mean +/- S.E.M. p values<0.05, calculated using unpaired student t-test. Data information: All invasion and migration assays were performed in triplicates and 6 different fields at 10x magnification were counted in 3 separate inserts. The data are presented as the mean +/- S.D. p values<0.05, calculated using unpaired student's t-test. B Quantification of transwell migration assays performed for PC3-LN4 EDC3 wild type, EDC3 S161A, EDC3 KO, and EDC3 S161D cell lines. The number of migrated cells (40 h) was counted as outlined in Materials and Methods. Data information: All invasion and migration assays were performed in triplicates and 6 different fields at 10x magnification were counted in 3 separate inserts. The data are presented as the mean +/- S.D. p values<0.05, calculated using unpaired student's t-test. C Quantification of transwell migration assays performed for PC3-LN4 EDC3 KO cells containing Doxycycline (DOX)-inducible EDC3-S161A with and without DOX (100ng/ml) induction. The number of migrated cells (40 h) was counted as outlined in Materials and Methods. Data information: All invasion and migration assays were performed in triplicates and 6 different fields at 10x magnification were counted in 3 separate inserts. The data are presented as the mean +/- S.D. p values<0.05, calculated using unpaired student's t-test. D Quantification of transwell invasion assay comparing PC3-LN4 EDC3 wild type, EDC3 S161A, EDC3 KO, and EDC3 S161D cell lines. Cells were plated in trans-wells coated with Matrigel and incubated for 72 h. The cells that invaded the matrix were counted as outlined in Materials and Methods. Data information: All invasion and migration assays were performed in triplicates and 6 different fields at 10x magnification were counted in 3 separate inserts. The data are presented as the mean +/- S.D. p values<0.05, calculated using unpaired student's t-test. E Quantification of transwell invasion assay performed for PC3-LN4 EDC3 KO cells containing DOX- inducible EDC3-S161A were either treated with or without DOX (100 ng/ml). The number of cells that invaded were counted as outlined in Materials and Methods. Data information: All invasion and migration assays were performed in triplicates and 6 different fields at 10x magnification were counted in 3 separate inserts. The data are presented as the mean +/- S.D. p values<0.05, calculated using unpaired student's t-test. F Relative mRNA expression of ITGA6, ITGB1, and KLF4 in PC3-LN4 WT and EDC3 S161A mutant cell lines. Data information: Real-time PCR data (F are presented as mean ± SEM, (n = 3/group). Unpaired student's t-test was used for statistical analysis. *P < 0.05, **P < 0.01 and NS, not significant. G Western blot analysis of proteins extracted from PC3-LN4 EDC3 wild type, EDC3 S161A, EDC3 knockout (KO, C60 cell line), and EDC3 S161D cell lines using the specified antibodies. Data information: All invasion and migration assays were performed in triplicates and 6 different fields at 10x magnification were counted in 3 separate inserts. The data are presented as the mean +/- S.D. p values<0.05, calculated using unpaired student's t-test. H Relative mRNA expression of ITGA6, ITGB1, and KLF4 in PC3-LN4 EDC3 KO-C60 cells, containing reexpressed EDC3 WT or EDC3 S161A mutant cDNAs Data information: Real-time PCR data I Western blot analysis of proteins extracted from PC3-LN4-EDC3-KO cells expressing GFP, EDC3 WT or the EDC3 S161A mutant using the specified antibodies. Data information: All invasion and migration assays were performed in triplicates and 6 different fields at 10x magnification were counted in 3 separate inserts. The data are presented as the mean +/- S.D. p values<0.05, calculated using unpaired student's t-test. J Western blot analysis of proteins extracted from PC3-LN4 cells treated with DMSO or the combination of Pim447 and AZD5363 for 96 h using the specified antibodies.