Fig. 3

The effect of N-BLR knockdown on invasion by specific siRNAs. a N-BLR abundance is decreased in stably silenced clones. b Invasion assays at 36 h show significant reduction of stably silenced N-BLR invading cells. c Migration assay at 24 h identified also significant reduction in migration of stably silenced N-BLR clones. d The 12 most significantly differentially expressed genes for both upregulated and downregulated genes. The data originated from 44 K Agilent microarray where HCT116 stable shRNA N-BLR clones #3-1 and #4-7 were compared with HCT116 empty vector control clone. The probes recognizing E-cadherin and vimentin are in red and blue, respectively. e Confirmation of microarray data by real time PCR shows that E-cadherin is increased and vimentin is markedly decreased in stably silenced clones (#3-1 and #4-7). f E-cadherin, vimentin, and ZEB1 were identified in vitro by immunofluorescence with specific antibodies. Immunofluorescence signal of E-cadherin (green color) was markedly increased in both clones. The ZEB1 signal was present in cells with empty vector (green color) but not in clones #3-1 and #4-7. Blue color indicate nuclei. Single green, blue, and merged channel images of ZEB1 are reported in Additional file 3: Figure S9B. g ZEB1 mRNA downregulation in HCT116 stable shRNA N-BLR clones #3-1 and #4-7 compared with control HCT116 empty vector clone. h Western blotting for E-cadherin and ZEB1 measured in the same clones; vinculin was used as loading control. (Student’s t-test; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001)