Fig. 4.

Ad19a E3/49K interacts with high molecular weight protein species in the T lymphocyte cell line Jurkat, but not in the epithelial cell line A549. Then 2 × 10⁶ A549 (lanes 1–6) or 4 × 10⁶ Jurkat cells (lanes 7–9) cells were mock-infected (−) or infected at a multiplicity of infection of 10 with Ad19a (WT) or an Ad19a mutant (49K KO), in which 49K expression was specifically abolished by inducing a frameshift (12). Cells were labeled 6 h and 18 h postinfection respectively, for 1.5 h and then lysed. In some cases (lanes 4–6), 0.5 mL of Jurkat lysate was added to 0.5 mL of A549 lysates as indicated, whereas in other samples (lanes 1–3 and 7–9), 0.5 mL of lysis buffer was added instead. For coprecipitation with purified sec49K, 600 ng of purified sec49K was added, followed by incubation for 1 h at 4 °C before immunoprecipitation. Immunoprecipitation was performed with antibodies directed against the C terminus (lanes 1–9) or N terminus (lanes 10–13). Arrows indicate full-length E3/49K (49K) and E3/49K-derived species, and arrowheads indicate the three protein species specifically binding to E3/49K in Jurkat cell lysates.