Fig. 3. Substitution of the dileucine motif of 5-HT_1AR results in ER sequestration. (A) Transfection of COS-7 cells with Flag-5-HT_1A (wild type), Flag-I414/415A-5-HT_1A or Flag-C417/420S-5-HT_1A. Cells were permeabilized and anti-Flag labeling is shown in green and anti-calregulin or anti-giantin labeling is shown in red. Bar, 10 µm. (B) Glycosylation state of receptors was analyzed by western blotting of crude membranes from transfected COS-7 cells with anti-5-HT_1A polyclonal antibody. Membranes were treated with Endo H, PNGaseF or untreated. Similar results were obtained in three independent experiments.