FIGURE 4.

APJ-S348A mutation impairs agonist-induced receptor internalization in HEK293 cells. HEK293 cells were co-transfected with β-arrestin-EGFP and the HA-tagged APJ construct at 1:1 ratio over the ensuing time period with apelin-13 (100 nm) for 0, 15, 30, and 60 min. The redistribution and co-localization of HA-APJ (red) and β-arrestin1-EGFP (A) or β-arrestin2-EGFP (B) after agonist stimulation were shown. The redistribution and co-localization of the HA-APJ-S348A (red) and β-arrestin1-EGFP (C) or β-arrestin2-EGFP (D) after agonist stimulation were shown. The nuclei were stained with DAPI. Yellow indicates the co-localization of β-arrestin and receptor. The results shown are representative images of at least three independent experiments. Vertical arrows show plasma membrane, slanted arrows show endosomes. Scale bars, 10 μm.