Figure 3.

SK2 is sufficient to promote ERM activation. A) HeLa cells were transfected with mock or SK2 DNA for 24 h. Cells were then starved for 4 h prior to stimulation with EGF for 30 s. Cells were then lysed, and SK2 activity was measured using an SK2-specific activity assay as described in the Materials and Methods. B) HeLa cells were transfected with vector, WT SK2, and G213E SK2 DNA for 24 h. Cells were then starved for 4 h prior to treatment with EGF (10 ng/ml) for 5 min. pERM and total SK2 levels were then assessed by immunoblotting. Total ezrin and actin were also included as loading controls. C) Quantification of the ratio of pERM:total ezrin in (D) was performed using ImageJ software. The data represent means ± se of 3 independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001. D) HeLa cells were transfected with vector, WT SK2, Ser387D;Tyr614D SK2, and Ser387A;Tyr614A SK2 DNA for 24 h. Cells were then starved for 4 h prior to treatment with EGF (10 ng/ml) for 5 min. pERM and total SK2 levels were then assessed by immunoblotting. Total ezrin and actin were also included as loading controls.