SHP2 rather than Src family kinases is involved in Erk activation and proliferation/survival upon FL stimulation in 32D cells. (A) Ten micromolar PP1, 15 μM PP2, or DMSO was added to Flt3-32D cells 15 minutes before ligand stimulation. FL (100 ng/mL) was added for the indicated periods of time before cells were lysed and cell lysates were subjected to a Western blot analysis for pErk and total Erk2. (B) SHP2 siRNA was introduced into WT- and Y599F-Flt3-32D cells by electroporation 72 hours before the experiment was performed. SHP2 knockdown (KD) was confirmed by probing total cell lysates for SHP2 and Erk2, respectively. Bands were densitometrically quantified and relative SHP2/Erk2 ratios are depicted under each lane (i). Parental and SHP2-KD cells were starved and stimulated with FL (100 ng/mL) as indicated before lysates were subjected to Western blot analysis for pErk and Erk2, respectively (ii). Densitometric values for the relative pErk/Erk ratios are depicted in 5Biii. (C) WT-Flt3-32D cells, Y599F-32D-Flt3, and KD counterparts were assessed for FL-dependent proliferation/survival by an MTT assay. Data from 3 individual experiments were subjected to ANOVA statistical analysis. Error bars indicate standard deviation. ^*P < .05; ^**P < .01; ^***P < .001.