FIG 9.

ASFV MGF110-7L induces ER stress. (A) 3D4/21 cells were transfected with a MGF110-7L-Flag-expressing vector with an increasing dose (0.5, 1.0, and 2.0 μg) or an empty Flag vector (2.0 μg) for 24 h. A positive control for ER stress was set by TG (1 μM) treatment for 6 h. Whole-cell lysates were obtained and immunolabeled with the indicated antibodies. Densitometry analysis of these protein bands in MGF110-7L-transfected cells normalized to empty vector-transfected cells was performed by ImageJ (right). (B to H) 3D4/21 cells were treated as described above for panel A. RNA samples were extracted at the indicated times, and the mRNA levels of BIP (B), XBP1(s)/XBP1(u) (C), EDEM1 (D), P58IPK (E), ERdj4 (F), GRP94 (G), and Calreticulin (H) were determined by RT-qPCR analysis. (I) Representative immunofluorescence images of PK-15 cells transfected with MGF110-7L-Flag-expressing vector (0.5 μg) for 24 h or treated with TG (1 μM) for 6 h and then incubated with anti-ATF6 (green) and anti-FLAG (red) antibodies. Nuclei were counterstained with DAPI (blue). Scale bar, 20 μm. Data in panels A to H show means ± the SD of the results of three independent experiments. *, P < 0.05; **, P < 0.01; ns, not significant.