Fig. 8

ARNTL2/E2F1 axis-mediated glycolysis sensitizes PC cells to erlotinib treatment via activating the PI3K/AKT pathway. A Volcano plot of differentially expressed gene profiles (ARNTL2-Knockdown vs. ARNTL2-Vector); B Heatmap of the indicated genes in PANC-1 cells with or without ARNTL2 knockdown; C KEGG analysis indicated that PI3K-Akt signaling pathway served as one of the major enriched signaling in ARNTL2-Knockdown group; D The mRNA levels of the indicated genes in PANC-1 cells with or without ARNTL2 knockdown; E GSEA analysis identified the glycolytic status in high ARNTL2 expression group; F Western blot analysis identified that the protein levels of PI3K-Akt signaling pathway and glycolysis related molecules in PANC-1 cells were decreased in ARNTL2-silenced group; G Glycolysis rate of PANC-1 cells ARNTL2 knockdown was examined by a Seahorse XFe96 Glycolysis Stress Test analyzer; H Western blot analysis identified that the protein levels of PI3K-Akt signaling pathway and glycolysis related molecules in PATU-8988 T cells were increased in ARNTL2-overexpressed group; I Glycolysis rate of PATU-8988 T cells overexpressing ARNTL2 was examined by a Seahorse XFe96 Glycolysis Stress Test analyzer; J-K Venn diagrams of four gene lists: transcription factors predicted by the hTFtarget, CHEA, ENCODE and JASPAR databases, ARNTL2 is transcriptionally activated by E2F1; L E2F1-binding motifs and predicted E2F1-binding sites (E1and E2) on the promoter region of ARNTL2 were obtained from the JASPAR database; M Chromatins were isolated from PANC-1 and BxPC-3 cells. The binding of E2F1 and blank control (Water) to the ARNTL2 promoter was tested using ChIP assay. N and O PANC-1 and BxPC-3 cells were incubated with E2F1 shRNA-1 and shRNA-2 or control (NC), western blot and qRT-PCR were used to test the protein and mRNA levels of ARNTL2. P Schematic representation depicting the mechanisms that the E2F1/ARNTL2 mediated glycolysis sensitizes PC cells to erlotinib treatment via activating PI3K/AKT pathway. All data are presented as the mean ± SEM of triplicate experiments. *p < 0.05, **p < 0.01 by repeated measures with Student’s t-test