Identification of the AKAP450 cis-Golgi–targeting domain by serial truncations. (A and D) Schematic representation of the AKAP450-truncated mutants used in this study. (right) A summary of the subcellular localization of each fragment. (B, C, and E) GFP-AK1 (B)–, GFP-AK4 (C)–, or GFP-AK1B (E)–expressing cells were labeled for GMAP210 (B and E) or γ-tubulin (C). Merged images are shown on the right. (F) WB of RPE-1 cells expressing AKAP450-truncated mutants used in this study. Molecular masses are given in kilodaltons. (G) Box and whisker plot showing quantification of GA recruitment for each N-terminal mutant. Top and bottom ends of the boxes represent 75th and 25th percentiles, and whiskers represent 90th and 10th percentiles. The median is depicted with a solid line. Asterisks denote significant statistical differences (P < 0.001; Tukey honestly significant difference [HSD]). n = 30 cells scored per mutant from three independent experiments. AU, arbitrary unit. (H) Merged images of cells expressing flag-AK1 or flag-AK1B double labeled for flag and either GM130 or Golgin-245. Fluorescence intensity profiles corresponding to lines drawn in each image from the top gallery are shown on the bottom. PACT, pericentrin-AKAP450 centrosomal targeting domain; WT, wild type. Bars, 5 µm.