LTK regulates ER export. (A) Representative images of HeLa cells stably expressing the GFP-RUSH-Man-II construct (Str-KDEL-Man-II-EGFP) under different conditions: 0 min, cells not treated with biotin; 20 min, cells fixed 20 min after biotin treatment; Cont, control siRNA transfected; siLTK, LTK silenced; crizotinib and alectinib indicate cells treated with 1 µM 30 min before biotin addition. Bar graph shows quantification from three independent experiments. Asterisk indicates statistically significant differences at P < 0.001. Scale bars in this figure are 15 µm. (B) Representative images of HeLa cells stably expressing the GFP-RUSH-Man-II construct (Str-KDEL-Man-II-EGFP) imaged 2 h after release of the reporter from the ER. Two conditions are depicted, control and LTK knockdown cells. (C) HeLa cells expressing the RUSH-Collagen-X construct were transfected with control or LTK siRNA. After 72 h, cells were treated with biotin and fixed immediately (T0) or after 20 min. Cells were immunostained against Giantin to label the Golgi. The increase in green fluorescence in the Golgi region relative to outside the Golgi region was measured using ImageJ. The bar graph represents the mean of four independent experiments.