Mutations in the EBMs strongly reduce the interaction of SMG6 with PABP. (A) V5-tagged SMG6 wild type or the indicated EBM mutants were expressed in human cells and immunoprecipitated using anti-V5 antibodies. The presence of endogenous PABP, Y14, and Magoh in the immunoprecipitates was analyzed by Western blotting. (B) UPF3b competes with SMG6 for binding to the EJC. GST pull-downs of the preassembled EJC with GST-UPF3bΔN or GST-SMG6 (1–207). (Lanes 1,2,5,6) To test for competition of EJC binding, the pull-downs with GST-UPF3bΔN were done in the absence or presence of untagged SMG6 (1–207). (Lanes 3,4,7,8) Conversely, the pull-downs with GST-SMG6 (1–207) were done in the absence or presence of MBP-UPF3bΔN. Samples were analyzed as described in Figure 4.