Fig. 3.

SG-like structures formation in EV71 infected cells is dependent on PKR and requires ongoing cellular mRNA synthesis. (A) RD cells were mock-infected or infected with EV71 (MOI = 10) for the indicated time, the cells were then lysed and Western blot was performed with the indicated antibodies. (B) RD cells were transfected with corresponding shRNA for 48 h and then infected with EV71 for indicated time, after which Western blot was performed with the indicated antibodies. (C) The CON^kd and PKR^kd cells were mock-infected or infected with EV71 for 7 h, after which the induction of SG-like structures was quantified by counting the number of foci in each cell within three independent immunofluorescence images that each contained more than 20 cells. PKR knockdown efficiency was confirmed by Western blot. (D) Blank cells and cells treated with 6 mM 2-AP for 7 h were mock-infected or infected with EV71, after which formation of SG-like structures was quantified as described in (C) and cell lysates were subjected to Western blot with the indicated antibodies. (E) RD cells were transfected with empty vector or vector encoding eIF2α(S52A) for 36 h, then infected with EV71 for 7 h and formation of SG-like structures was quantified as described in (C) and cell lysates were subjected to Western blot with the indicated antibodies. (F) RD cells were mock-infected or infected with EV71 for 7 h and ActD was added at 3 h post-infection. The cells were then fixed and stained for HuR and nuclei.