c-MplK553+573R exhibits altered Tpo-induced proliferation and degradation. (A) MTT proliferation assay using BaF cells expressing WT c-Mpl or c-Mpl553+573R. Cells were treated with rhTpo at increasing concentrations for up to 48 hours. The data represent the mean (± SE) of 3 individual experiments using 2 stably expressing clones for each group (*P < .05, ***P < .001). (B) Western blot analysis of Tpo-induced c-Mpl phosphorylation with and without MG-132. The data are representative of 3 individual experiments. (C) Western blot analysis of Tpo-induced c-Mpl degradation of WT c-Mpl and c-MplK553+573R. Cells were pretreated with Chx for 60 minutes before Tpo stimulation for up to 120 minutes. (D) Graphic representation of c-Mpl degradation, comparing mature (85-kDa) c-Mpl at time 0 using densitometry.
