FIG 7.

eIF4B S406 phosphorylation positively regulates the MET expression. (A and B) U2OS cells were transfected with a control vector or plasmids expressing wild-type eIF4B (WT) and its mutants for 48 h. Cell lysates were analyzed by immunoblot assays using indicated antibodies. (C) U2OS cells were transfected as in panels A and B. At 24 h after transfection, cells were treated with AZD1208 (3 μM) for an additional 24 h. Cell lysates were analyzed by immunoblot assays using indicated antibodies. (D and E) U2OS cells overexpressing eIF4B or its mutants S406A, S422A, S406/422A, S406D, and S406E were labeled with ³⁵S for new protein synthesis. Newly synthesized MET and ERK were immunoprecipitated, separated by SDS-PAGE, and visualized by autoradiography. (F) HeLa cells were transfected with pR-MET-F construct together with eIF4B or its mutants. After 24 h, luciferase assays were performed. Relative ratios of firefly/Renilla luciferase activities are shown. The ratio for the vector control was set as 1. The averages ± SDs are shown. P < 0.05, WT versus vector; P < 0.02, WT versus S406A or S406/422A.