FIG. 3.

MAPKKs as mediators of Mxi2 activation. (A) Effects of MAPKK dominant inhibitory mutant forms on Mxi2 activation induced by EGF. Cells were transfected with AU5-tagged Mxi2 and the indicated HA-tagged inhibitory mutant forms (250 ng). The indicated cells were stimulated with EGF (100 ng/ml) for 5 min. The data shown are the average ± the standard error of the mean of three independent experiments relative to the activity detected in control cells. (Right side) Expression levels of the MAPKKs in total lysates. (B) Activation of Mxi2 by MAPKKs. Kinase activities in cells transfected with HA-tagged Mxi2 or p38, together with the indicated MAPKKs (1 μg). (Bottom) Expression levels of HA-Mxi2 and of the different MAPKKs. (C) Activation of Mxi2 by constitutively active MEK isoforms. Kinase activities detected in cells transfected with AU5-Mxi2 and activated, HA-tagged MEK1 and -2 (1 μg) or stimulated with EGF. (Bottom) Expression levels of AU5-Mxi2 and HA-MEK1 and -2. WB, Western blot; c, control; v, vector.