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Figure 5 | Journal of Hematology & Oncology

Figure 5

From: Long non-coding RNA ANRIL is upregulated in hepatocellular carcinoma and regulates cell apoptosis by epigenetic silencing of KLF2

Figure 5

ANRIL could silence KLF2 expression. (A) Co-expression analysis by using GSE45435 data from GEO datasets. (B) The KLF2 gene expression in HCC by using GSE 56140. (C-E) The levels of KLF2 mRNA and protein were detected by qPCR and Western blot when HepG2 and Hep3B cells were transfected with si-ANRIL, and results are expressed relative to the corresponding values for control cells. (F-H) The levels of KLF2 mRNA and protein were detected by qPCR and Western blot when HepG2 and Hep3B cells were transfected with si-EZH2, and results are expressed relative to the corresponding values for control cells. (I-K) The levels of KLF2 mRNA and protein were detected by qPCR and Western blot when HepG2 and Hep3B cells were transfected with si-SUZ12, and results are expressed relative to the corresponding values for control cells. (L,M) ANRIL expression levels in cell cytoplasm or nucleus of HCC cell lines Hep3B and HepG2 were detected by qPCR. (N,O) 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 ANRIL 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. (P,Q) 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 ANRIL siRNA (48 h) or scrambled siRNA. (R) The KLF2 expression level was determined by qPCR in mice tumors formed from HepG2/sh-ANRIL and HepG2/empty vector. (S) Tumors developed from sh-ANRIL-transfected HepG2 cells showed higher KLF2 protein levels than tumors developed by control cells. **P < 0.01.

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