Figure 4

ERK1/2 activation by FLT3 ITD in MV4-11 cells is negatively regulated by DUSP6. MV4-11 cells were transiently transfected with DUSP6-targeting siRNA by nucleofection. After 48 h, cells were serum-starved for 4 h and then stimulated with 100 ng/ml FL for the indicated times. Cells were extracted and subjected to immunoblotting for detection of ERK1/2 phosphorylation, and DUSP6 levels. ERK1 was assessed as loading control. (A) Representative experiment. Unstimulated cells, which were cultivated in complete medium, were also analyzed (as indicated). The data for the latter are from the same blot with identical exposure and image processing, but were rearranged for better clarity. (B) Quantification of immunoblots for 4 independent experiments (except time-point 10 min, n = 2). Statistic siginificance was tested by Students’ test (*p < 0.05) for DUSP6 siRNA versus control. (C) Effect of FL stimulation on DUSP6 protein levels. Quantification of immunoblots for 4 independent experiments. DUSP6 signals were normalized to ERK1. Statistic significance was tested by Students’ test (*p < 0.05,**p < 0.01) for difference from unstimulated control.