FIGURE 3.

Suppression of PKR expression by shRNA suppresses B56α expression and promotes BCL2 phosphorylation. A, Western blot analysis was performed using antibodies against PKR, B56α, p-eIF2 α, eIF2 α, BCL2, and actin on total lysate (1 × 10⁶ cell equivalents) from REH cells transfected with control shRNA or PKR shRNA. B, REH cells transfected with control shRNA or REH cells transfected with PKR shRNA were labeled with [³²P]orthophosphoric acid (³²P). BCL2 was immunoprecipitated using polyclonal rabbit antisera (Santa Cruz Biotechnology), electrophoresed using 10% SDS-PAGE, and transferred to a nitrocellulose filter, and phosphorylated bands were detected by autoradiography. The identity of the BCL2 ³²P-labeled band was established by using a mouse monoclonal antibody against BCL2 (Dako) on the same filter. C, Western blot analysis was performed using antibodies against B55, PP2A/A, and GAPDH on total lysate (1 × 10⁶ cell equivalents) from REH cells transfected with control shRNA or PKR shRNA.