Fig. 4

TUG1 could silence KLF2 expression. a KLF2 gene expression in HCC tissues (n = 77) compared with corresponding non-tumor tissues (n = 77). KLF2 expression was examined by qPCR and normalized to GAPDH expression. Results were presented as ΔCT in tumor tissues relative to normal tissues. b Co-expression analysis between TUG1 and KLF2. c The levels of KLF2 mRNA were detected by qPCR when HepG2 and Hep3B cells transfected with si-TUG1 and results are expressed relative to the corresponding values for control cells. d,e The levels of KLF2 protein levels were determined by Western Blotting when HepG2 cells transfected with si-TUG1. f,g The levels of EZH2 protein were detected by Western Blotting when HepG2 and Hep3B cells transfected with si-EZH2 and results are expressed relative to the corresponding values for control cells. h The levels of KLF2 mRNA were detected by qPCR when HepG2 and Hep3B cells transfected with si-EZH2 and results are expressed relative to the corresponding values for control cells. i,j The levels of EZH2 protein were detected by Western Blotting when HepG2 and Hep3B cells transfected with si-EZH2 and results are expressed relative to the corresponding values for control cells. k,l The levels of SUZ12 protein levels were determined by Western Blotting when HepG2 cells transfected with si-SUZ12. m The levels of KLF2 mRNA were detected by qPCR when HepG2 and Hep3B cells transfected with si-SUZ12 and results are expressed relative to the corresponding values for control cells. n,o The levels of KLF2 protein levels were determined by Western Blotting when HepG2 cells transfected with si-SUZ12. p,q TUG1 expression levels in cell cytoplasm or nucleus of HCC cell lines Hep3B and HepG2 were detected by qPCR. r,s RIP with rabbit monoclonal anti-EZH2, anti-SUZ12, anti-SNRNP70 and preimmune IgG from HepG2 and Hep3B cell extracts. RNA levels in immunoprecipitates were determined by qPCR. Expression levels of TUG1 RNA were presented as fold enrichment in EZH2 and SUZ12 relative to IgG immunoprecipitates; relative RNA levels of U1 snRNA in SNRNP70 relative to IgG immunoprecipitates were used as positive control. t,u ChIP–qPCR of EZH2 occupancy and H3K27-3me binding in the KLF2 promoter in HepG2 cells, and IgG as a negative control; ChIP–qPCR of EZH2 occupancy and H3K27-3me binding in the KLF2 promoter in HepG2 cells transfected with TUG1 siRNA (48 h) or scrambled siRNA. v The KLF2 expression level was determined by qPCR in mice tumors formed from HepG2/sh-TUG1,HepG2/empty vector. w Tumors developed from sh-TUG1 transfected HepG2 cells showed higher KLF2 protein levels than tumors developed by control cells. *P < 0.05, **P < 0.01 and N.S. not significant