ER Stress is activated in BHK-21 cells transfected with 2C- or 3D-expressing plasmid. (A) A Western blotting analysis of GRP78, GRP94 and β-actin in the extracts of BHK-21 cells that were treated with 500 nM Tg for 12 h, infected with EMCV (MOI = 0.005) for 12 h or transfected with 2C- or 3D-expressing plasmid for 48 h. (B) The BHK-21 cells were transfected with pGL3-GRP78-luc, pGL3-GRP94-luc, pGL3-Carlreticulin-luc, pGL3-ATF4-luc or pGL3-CHOP-luc along with pRL-TK and pCMV-HA-2C, pCMV-HA-3D or pCMV-HA. The firefly luciferase activity was measured in cell lysates that were normalized with Renilla luciferase activities at 48 h post-transfection. Data are expressed as the means ± SD. **p < 0.01; ***p < 0.001, compared with the corresponding control. HA, the BHK-21 cells transfected with pCMV-HA and Mock, the uninfected or untransfected BHK-21 cells, as the control. (C) The conversion of LC3 by ER-stress inducer Tg. BHK-21 cells without treatment or with a 500 nM Tg treatment for 12 h were analyzed by Western blotting for LC3 and β-actin.