Figure 9.

(A, B) HEK-293 cells, transfected with the wild-type Mash1 with or without E47 expression constructs as indicated, were labeled with [³⁵S]methionine and treated for 2 h with the drugs indicated (FK (forskolin); H89; DRB (5,6-dichloro-1-β-ribofuranosylbenzimidazole); U (U0126); bisI (bisindoleylmaleimide I); SB (SB203580); emo (emodin); api (apigenin)). Immunoprecipitated Mash1 and co-immunoprecipitated E47 were visualized as described above. (C) HEK-293 cells were transfected with the indicated expression constructs and siRNA for CK2β subunit mRNA or an unrelated siRNA directed against uPF2K as a control. After 24 h, cells were either metabolically labeled and both Mash1 and co-immunoprecipitated E47 were visualized as described above, or lysed and both CK2β and tubulin were analyzed by immunoblotting. (D) DMS53 cells were treated with DRB, emodin or apigenin for 2 h and Mash1 or α-tubulin were visualized by immunoblotting. (E) Immunoprecipitates of extracts from HEK-293 cells transfected with the indicated constructs were split and either incubated with recombinant CK2 and [γ-³²P]ATP (upper panel) or subjected to immunoblotting in order to visualize the relative amounts of immunoprecipitated Mash1 (lower panel).