FIG. 2.
(A) Overexpression of RafΔ26–303-Cx leads to phosphorylation of endogenous ERK1/2. A3.01 T cells were transfected with either RafΔ26–303, RafΔ26–303-Cx the corresponding kinase-dead mutant (RafΔ26–303-KD-Cx), or empty expression vector as a control (mock). Unstimulated or TPA-stimulated (25 ng/ml for 20 min) T cells were lysed in RIPA buffer 42 h after transfection, proteins were separated by SDS-PAGE, and immunoblots were prepared with phospho-ERK specific antibodies. (B) Corresponding immunoblots obtained with anti-panERK antiserum to demonstrate that there were equal amounts of protein kinase.