Figure 2.

Five RGS4 mutant proteins do not inhibit G-protein-mediated MAPK activation. (A) Mammalian expression plasmids containing RGS4 or RGS4 mutant proteins L159F, S164Q, R167M, F168A, and N88S were transfected into 6E cells. (Upper) MAPK activity is represented by the amount of [γ³²P]ATP incorporated into myelin basic protein incubated with HA immunoprecipitates (MBP, above, labeled “kinase”). The average increase in activity over that of unstimulated cells as determined from three independent experiments is shown in the bar graph + SEM. (Lower) The levels of cotransfected HA-ERK1 as determined by immunoblotting the immunoprecipitates used to determine kinase activity with anti-HA (labeled “WB,” below). (B) Levels of indicated mutants were determined by immunoblotting equivalent amounts of cell lysates transfected with HA-tagged RGS4 expression plasmids with the anti-HA antibody.