FIG. 2.
MVMp gene expression and PKR activity in 3T3 fibroblasts. (A) Time course accumulation of MVMp proteins and eIF2α phosphorylation in wild-type 3T3 and PKRo/o cells. Infected cells were lysed in sample buffer and analyzed by Western blotting at the indicated time points. Once the anti-phospho eIF2α blot was carried out, the membrane was stripped and reprobed with anti-total eIF2α. (B) Protein synthesis in MVMp-infected 3T3 cells. Cells were pulse-labeled with [35S]Met (25 μCi/ml) for 30 min at the indicated time points (hpi), and labeled proteins were resolved by 10% SDS-PAGE and autoradiography. −, mock-infected culture. (C) IF analysis of eIF2α phosphorylation in MVMp-infected 3T3 cells. At 15 hpi, cells were fixed and simultaneously probed with anticapsid (red) and anti-phospho eIF2α (green). Arrows point to infected 3T3 cells showing some MVMp input viral particles accumulated in endosomal vesicles.