Figure 4

ESE-1 upregulated GP73 expression in HCC cells. (A) pCR3.1-ESE-1 plasmid was transfected into HEK293T cells and ESE-1 expression was tested by Western blot 36 h after transfection. (B) pCR3.1-ESE-1 plasmid was transfected into Hep3B cells for 36 h, and GP73 mRNA and protein levels were analyzed with qPCR and Western blot, respectively. (C) GP73 mRNA and protein levels were analyzed by adopting qPCR and Western blot in Huh7 cells transfected with pCR3.1-ESE-1 plasmid. (D) pCR3.1-ΔESE-1 plasmid with deletion ETS domain of ESE-1 was constructed and transfected into HEK293T cells. ΔESE-1 expression was confirmed by Western blot. (E) The empty vector (Mock) or ESE-1 or ΔESE-1 was transfected into Hep3B and Huh7 cells, and GP73 expression was tested by Western blot. (F) ESE-1 was downregulated in Hep3B and (G) Huh7 cells by infecting them with virus carrying the shRNA expression cassette against ESE-1 (shESE-1) or a non-target control (shGFP) with or without IL-1β stimulation. After 96 h, the mRNA and protein levels of ESE-1 and GP73 were analyzed with qPCR and Western blot, respectively. (H) Hep3B and (I) Huh7 cells infected with shESE-1 virus particles were transfected with pCR3.1-ESE-1 plasmid at 48 h post-infection. At 48 h after transfection, cells were harvested, and ESE-1 and GP73 proteins were analyzed with Western blot.