FIGURE 1.

ICP34.5 mediates the association of PP1 and eIF2α. A, fluorescence visualization and FACS analysis. HeLa cells were transfected with plasmids encoding YN-PP1 and YC-eIF2α, along with a plasmid encoding ICP34.5 (WT), ICP34.5 (V¹⁹³E, F¹⁹⁵L), or a control plasmid. Then cells were subjected to fluorescence microscopy (left column), and the FACS analysis profile of each transfectant was plotted in the center column, with the mean YFP fluorescence intensity (MFI) in the right column. B, the protein expression of each transfected component was verified by Western blot analysis with the indicated antibodies. C, coimmunoprecipitation (IP) of eIF2α with PP1. HEK293T cells were cotransfected with plasmids encoding FLAG-tagged YN-PP1 and YC-eIF2α together with pHIV-vFlip, pHIV-ICP34.5 (WT), or pHIV-ICP34.5 (V¹⁹³E, F¹⁹⁵L). At 48 h post-transfection, cell lysates were collected and incubated with anti-FLAG antibody and protein A/G-agarose for 2 h at 4 °C with gentle agitation. The immobilized proteins were resolved by Western blot analysis using antibodies against eIF2α and FLAG, respectively.