FIGURE 3.

RMB4-mediated translation suppression involves Ago2. A, cell lysates were prepared from FLAG-RBM4-expressing or mock C2C12 cells grown in GM or DM and then subjected to anti-FLAG immunoprecipitation (IP) followed by immunoblotting. The luciferase assay in B and C was performed using the mock pRL or pRL-2×CU reporter in HEK293 cells and as described under “Experimental Procedures”; cotransfected vectors of effector proteins or siRNA are as indicated. Bar scales show relative RL activity (see under “Experimental Procedures”). Ago2 knockdown efficiency was assessed by using immunoblotting. D, pRL-2×CU was cotransfected with effector expression vector(s) as indicated. Immunoprecipitation was performed 48 h after transfection; coprecipitates were analyzed by RT-PCR using primers specific for the RL-2×CU mRNA.