Adaptor SKAP-55 Binds p21ras Activating Exchange Factor RasGRP1 and Negatively Regulates the p21ras-ERK Pathway in T-Cells
Figure 2
SKAP-55 knock-down and knock-up T-cells show altered ERK activation.
Panel A: Upper panel: Reduction in SKAP-55 expression in SKAP-55KD cells. Z.21, Z.22 and Z.23 cell lines were derived from distinct SKAP55 shRNAs as described in Material and Methods. Panel B: Increased pERK in response to anti-CD3 in SKAP-55 KD cells. T8.1 WT and T8.1 SKAP-55 KD cells (Z.21, Z.22, Z.23) were either left untreated (lanes 1, 7,13, 19) or stimulated with anti-CD3 (10 µg/ml) (lanes 2–6; lanes 8–12; lanes 14–18; lanes 20–24). Upper panels: anti-pERK blotting; lower panels: anti-ERK blotting. Panel C: Reduction of ERK activation in DC27.10 cells. DC27.10 cells (lanes 1–4) and cells transfected with SKAP-55 (lanes 5–8) were either left untreated (lanes 1,5) or stimulated with anti-CD3 (5 µg/ml) for the indicated times (lanes 2–4 and 6–8). Upper panels: anti-pERK blotting; middle panels: anti-ERK blotting; lower panels: anti-SKAP-55 blotting.