HPIV3 viral RNA triggers SG formation.

(A and B) HeLa cells were transfected with the indicated RNA samples from mock-infected or HPIV3-infected MK2 cells, treated with pIC for 12 h or AS for 1 h. (A) Cells were immunostained for TIA-1 (green) and G3BP (red). Nuclei were stained with DAPI (blue). The white scale bar corresponds to 10 μm. (B) Cell lysates were analyzed via western blot using anti-phosphorylated PKR, anti-PKR, anti-phosphorylated eIF2α, anti-eIF2α, and anti-GAPDH antibodies. (C and D) HeLa cells were transfected with the RNA from HPIV3-infected MK2 cells, the respective PolyA⁺ RNA fraction, or the respective PolyA^- RNA fraction for 12 h. (C) Cells were immunostained for TIA-1 (green) and G3BP (red). Nuclei were stained with DAPI (blue). The white scale bar corresponds to 10 μm. (D) The percentage of cells containing SGs was quantified in three independent experiments. (E and F) HeLa cells were transfected with the respective PolyA⁺ RNA fraction from HPIV3-infected MK2 cells for 12 h and subsequently mock-treated or treated with CHX for another 1 h, 2 h, or 3 h. (E) Cells were immunostained for TIA-1 (green) and G3BP (red). Nuclei were stained with DAPI (blue). The white scale bar corresponds to 10 μm. (F) The percentage of cells containing SGs was quantified in three independent experiments. Data are represented as means ±SD. Student’s t test: * P<0.05, ** P<0.01, *** P<0.001, ns = not significant.