Fig. 3

EIF4A3 promotes the biogenesis of circTOLLIP. a Prediction of the putative EIF4A3 binding sites in TOLLIP pre-mRNA by CircInteractome database. b Six positions (a-f) in TOLLIP pre-mRNA were selected to design qPCR primers and 4 plasmids (A1-A4) containing EIF4A3 binding sites were constructed to pull down EIF4A3 protein. c RIP assay using EIF4A3 antibody verified the direct binding of EIF4A3 and TOLLIP pre-mRNA. d Western blot analysis of EIF4A3 protein with MS2-RNA pulldown assay. Vector and GAPDH were used as negative controls. e-f qRT–PCR analysis of circTOLLIP level in HLF and 97H cells with transfection of EIF4A3 siRNA or EIF4A3 overexpression plasmid. g-h Stability of circTOLLIP in EIF4A3-overexpressing or EIF4A3 knockdown 97H cells with actinomycin D treatment. i Representative images of IHC staining in 138 pairs of HCC tissues. j Statistical analysis of the IHC results. k Correlation analysis between circTOLLIP and EIF4A3 expression in HCC tumors