Rescue of SHP2 expression in TF-1 cells. (A). TF-1 cells transduced with empty vector (MIG-R1) as well as vectors carrying WT SHP2 cDNA (SHP2-WT) and SHP2 cDNA with noncoding mutations in the sh-Shp2-2 target sequence (SHP2-shR) were transduced with sh-Ctrla– and sh-Shp2-2–expressing vectors, respectively. GFP⁺DsRED⁺ cells were sorted by FACS and expression of SHP2 analyzed by Western blotting. SHP2-WT expression resulted in partial restoration of SHP2 expression in sh-Shp2-2 shRNA-expressing cells, whereas SHP2-shR completely rescued expression of SHP2 to normal or even higher levels in sh-Shp2-2 shRNA-expressing cells. (B) Dual-transduced TF-1 cells were cultured with GM-CSF (2 ng/mL). Apoptosis was analyzed by labeling with annexin V and DAPI. Data are mean ± SEM of fold changes of apoptosis from sh-Shp2-2 over sh-Ctrla-expressing cells from 3 experiments. *P < .05 for SHP2-WT compared with SHP2-shR, and P < .01 for MIG-R1 compared with SHP2-WT and SHP2-shR. (C) Dual-transduced TF-1 cells were starved of GM-CSF overnight and then cultured with GM-CSF (0.3 ng/mL). Cell growth in response to the GM-CSF was evaluated by MTS assay. Inhibition of cell growth of sh-Shp2-2–transduced cells relative to the cell growth of the corresponding sh-Ctrla–transduced cells was calculated. Data are mean ± SEM from 3 replicates. *P < .05 for SHP2-WT compared with SHP2-shR, and **P < .01 for MIG-R1 compared with SHP2-shR.