Fig. 4.

Evaluation of the DN effect of the E555 variants. (A) E47 transcriptional activity after co-transfection with the expression vector containing E47 wild-type (WT) (1.5 ng) and each E555 variant (1.5, 3.0, or 4.5 ng). (B) Immunoblotting for subcellular E47 protein expression in human embryonic kidney (HEK) 293T cells co-transfected with WT and E555 variants. (C) Immunoblot analysis of E47 protein in total protein extracts from HEK293T cells co-transfected with WT and E555 variants (left). Whole-cell lysates were immunoprecipitated with anti-V5 antibody followed by immunoblotting with anti-FLAG or anti-V5 antibody (right). Immunoprecipitation with non-specific IgG was used as an experimental negative control. (B, C) Detection of E555* was weaker than in Fig. 3A due to the use of different primary antibodies (anti-human E47 or anti-FLAG antibody) (Fig. S5)