Fig 1: C9orf16 knockdown inhibited cancer cell invasion and increased apoptosis. A Expression of C9orf16 in pancreatic cancer cell lines and normal epithelial cells was examined by RT-PCR. B RT-PCR examination of C9orf16 knockdown cells and the scrambled cells showed the C9orf16 knockdown efficiency. C Real time PCR was used to check the C9orf16 knockdown efficiency. D-E Western blotting was performed to confirm the C9orf16 knockdown efficiency. F Cell proliferation measured by MTT assay showed decreased cell proliferation in PANC-1 cells with oeC9orf16 knockdown. Migration and invasion assays (G) and cell number quantification (H) of the C9orf16 knockdown and the scrambled cells. Flow cytometry analysis (I) of Cisplatin induced cell apoptosis and quantification of late apoptotic cell percentage (J) of the C9orf16 knockdown and the scrambled cells. Scale bar: 100 µm. **: p < 0.01; ***: p < 0.001; ****: p < 0.0001
Fig 2: C9orf16 overexpression in normal pancreatic epithelial cell increases cell invasion and decreased apoptosis. A-D RT-PCR (A), real time PCR (B) and western blotting (C-D) analysis were performed to confirm the overexpression efficiency. E Cell proliferation measured by MTT assay showed increased cell proliferation in HPNE cells with oeC9orf16 overexpression. Migration and invasion assays (F) and cell number quantification (G) of the C9orf16 overexpression and the scrambled cells. Flow cytometry analysis (H) of Cisplatin induced cell apoptosis and quantification of late apoptotic cell percentage (I) of the C9orf16 overexpression and the scrambled cells. Scale bar: 100 µm. **: p < 0.01; ***: p < 0.001; ****: p < 0.0001
Fig 3: Activated EMT in PDAC cancer cells. A Heatmap visualization of the expression of epithelial cell marker genes and mesenchymal signature genes in normal epithelial cells, primary and metastatic cancer cells by “DoHeatmap” function in R-studio (Version 1.4.1717). Relative expression of the epithelial cell gene, CDH1, and mesenchymal cell gene, VIM, in C9orf16 knockdown cancer cells (B) and C9orf16 overexpression normal epithelial cells (C). *: p < 0.05; **: p < 0.01; ***: p < 0.001
Fig 4: Pathway analysis revealed MYC signaling pathways as the top activated pathways. A Heatmap of the expression of the differentially expressed genes between C9orf16 knockdown and scrambled cancer cells by “DoHeatmap” function in R-studio (Version 1.4.1717). B Pathway analysis on differentially expressed genes of C9orf16 expression (scrambled) cancer cells relative to C9orf16 knockdown cells. C Pathway analysis on the differentially expressed genes of PDAC cancer cells relative to normal epithelial cells in the scRNA-seq data
Fig 5: Blocking MYC signaling reduced cancer cell invasion and increased apoptosis. A MYRA-A and KSI-3716 treatment effectively blocked the activity of Myc signaling pathway in PANC-1 cells. B Real time PCR analysis of C9orf16 expression after MYRA-A and KSI-3716 treatment. C-D Western blotting analysis were performed to examine the expression of C9orf16 after the treatment of MYRA-A and KSI-3716 as well as recombinant human c-Myc protein. Migration and invasion assays (E) and cell number quantification (F) on cancer cells after MYRA-A and KSI-3716 treatment. Flow cytometry analysis (G) of Cisplatin induced cell apoptosis and percentage quantification of late apoptotic cells (H) of cancer cells after MYRA-A and KSI-3716 treatment. Scale bar: 100 µm. *: p < 0.05; **: p < 0.01; ***: p < 0.001; ****: p < 0.0001
Supplier Page from OriGene Technologies for BBLN Human qPCR Primer Pair (NM_024112)