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. 2004 May;24(10):4502–4512. doi: 10.1128/MCB.24.10.4502-4512.2004

FIG. 2.

FIG. 2.

PKR interacts with TRAF family proteins in vivo. (A) HeLa cells were infected with 10 PFU of VT7 per cell. After 1 h, plasmids encoding FLAG-tagged TRAF5 and TRAF6 proteins or the empty vector were transfected together with the empty vector or a plasmid encoding PKR (K296R mutant form). Cell extracts were collected 20 hpi and analyzed by SDS-PAGE, followed by immunoblot (Western blot [WB]) analysis with anti-PKR or anti-FLAG antiserum. (B) Extracts prepared as described for panel A were immunoprecipitated with anti-PKR serum and thoroughly washed, and immunocomplexes were analyzed by SDS-PAGE and subjected to immunoblotting with antiserum to PKR or FLAG. (C) HeLa cells in 10-cm-diameter plates were infected with 10 PFU of VT7 per cell. After 1 h, a plasmid (10 μg) encoding HA-tagged TRAF2 was transfected together with 10 μg of the empty vector (lane 1) or with 10 μg of a plasmid encoding PKR (K296R mutant form, lane 2). Cells extracts were collected at 20 hpi and analyzed by SDS-PAGE, followed by immunoblot analysis with anti-PKR or anti-HA antiserum. (D) Extracts prepared as described for panel C were immunoprecipitated with anti-HA serum and thoroughly washed, and immunocomplexes were analyzed by SDS-PAGE and subjected to immunoblotting with antiserum to PKR or HA. The asterisk denotes IgGs, and the arrows indicate the specific proteins.