Figure 4

CD157 promotes survival of leukemic blasts through Mcl-1 upregulation. (A) CD157-high (red line) or CD157-low (blue line) U937 cells were treated for 24 h with increasing concentrations of S63845, then cell viability was determined by Presto Blue assays. Data represents the percentage of live cells compared to control cells treated with vehicle and are the mean ± SEM of three experiments performed in six replicates. (B) Western blot analysis of PARP-1, Mcl-1 and cleaved Caspase-3 in CD157-high or CD157-low U937 cells treated with S63845 for 24, 48 or 72 h. β-Tubulin was used as loading control. Uncropped blot images are provided in Supplementary Fig. S11. (C) CD157-high or CD157-low U937 cells were treated for up to 72 h with S63845. Cell apoptosis was measured by flow cytometry analysis after AnnexinV/PI staining. Results are the mean ± SEM of three experiments. *p < 0.05, two-way ANOVA with Sidak’s multiple comparison test. (D) U937/CD157-high or U937/CD157-low cells were treated with S63845 for 24 h, then lysed and subjected to immunoprecipitation with anti-Mcl-1 or anti-Bim antibody, respectively. The interactions with the indicated proteins were analyzed by immunoblotting. β-Tubulin was used as loading control. One representative Western blot is shown (n = 3). The whole cell lysates (WL) show the protein expression levels. Uncropped blot images are provided in Supplementary Fig. S11.