Ei24 can inhibit IMPβ1- or IMPα2/β1-mediated nuclear accumulation dependent on a polyarginine sequence within its IBBL domain. (A) Live-cell CLSM images of HeLa-Bcl_XL cells transfected to coexpress the indicated GFP and DsRed2 fusion proteins 20 h after transfection. Bars, 10 µm. (B) Quantitative analysis for the extent of nuclear accumulation (Fn/c) of the various GFP fusion proteins. Results are for the mean ± SEM (error bars; n ≥ 41) from a single assay representative of three separate experiments; p-values denote significant differences. (C) Lysates from HEK293T cells transfected to express FLAG-Ei24 WT, RRRRm mutant derivative, or vector alone were subjected to immunoprecipitation using anti-FLAG M2 antibody. Western analysis was performed on input and immunoprecipitates (IP: FLAG) using specific antibodies against IMPβ1 or Ei24, with β-actin as a control. Densitometric analysis was performed on images such as those shown in C; results are for the mean ± SEM (error bars n = 3) for binding (%) relative to that for WT. (D) CLSM images of HeLa-Bcl_XL cells transfected to coexpress the indicated GFP fusion proteins in the absence or presence of FLAG-Ei24 derivatives were fixed 20 h after transfection before being immunostained using an anti-FLAG M2 antibody. Bars, 10 µm. (E) Quantitative analysis of the extent of nuclear accumulation for the various GFP fusion proteins. Results shown are for the mean ± SEM (error bars; n ≥ 40) for a single assay representative of three independent experiments. Statistical analysis was performed as in B.