AK1 and AK1B fragments differentially affect GA positioning and morphology. (A) RPE-1 cells expressing the flag-AK1 fragment were triple labeled for flag, AKAP450, and GMAP210. Single labelings and the merge are shown. White arrows and arrowheads indicate the GA and the CTR, respectively. In B, a nontransfected cell is shown for comparison. N, nucleus. (C) Merged image of a flag-AK1–transfected cell stained for flag, α-tubulin, and GMAP210 to visualize the MT network. (D) High magnification images of a flag-AK1–transfected cell triple labeled as in C. Single labelings in black and white and the merge are shown. Yellow arrows indicate the alignment of AK1B-containing structures with MTs. (E and F) Merged images of flag-AK1B–expressing cells triple labeled as in A or C, respectively. (G) Low magnification images of flag-AK1 (top)– and flag-AK1B (bottom)–transfected cells triple labeled for flag, AKAP450, and GMAP210. Single labelings for flag are shown on the left to identify transfected cells (T). (right) Schematic of the measurement of CTR–GA distance and GA diameter over the corresponding AKAP450 and GMAP210 merged images. A straight line connecting the center of the circumferences circumscribing the GA and the CTR was measured to calculate the CTR–GA distance. GA diameter was used as an index of GA circularity. NT, not transfected. (H) Graphs showing quantification of the CTR–GA distance (top) and Golgi circularity index (bottom; n = 160 for each condition from three independent experiments). Data are presented in the same box and whisker format as in Fig 1 with the median marked as a solid line (*, P < 0.001; Tukey HSD). Top and bottom ends of the boxes represent 75th and 25th percentiles, and whiskers represent 90th and 10th percentiles. Bars, 2.5 µm.