Skip to main content
. Author manuscript; available in PMC: 2020 Dec 15.
Published in final edited form as: Neuropharmacology. 2019 Jun 20;161:107676. doi: 10.1016/j.neuropharm.2019.107676

Fig. 6. Examples of the effects of mutations listed in Table 1 on localization and oligomerization of YFP-HA-DAT.

Fig. 6

(A) wt DAT (WT), L450/454/458A (LLL), C306A or W238/256A mutants were stably expressed as clonal pools in PAE cells (approximately 10–20% of cells are expressing DAT or its mutants). The cells were incubated with DMSO (vehicle; “-” AIM-100) or 20 μM AIM-100 for 2 hours at 37°C and lysed. Western blotting of cell lysates was performed using GFP antibody. hO, higher oligomers. All lanes are from the same blot. *Non-specific bands (detected due to low levels of expression of DAT and its mutants). The results are representative of 2–3 experiments with each mutant. (B-C) wt DAT (WT) and mutants H223A, D231/232A, or D231A/D232A/P235A/P236A/R237A (DDPPR) were transiently expressed in PAE cells. The cells were fixed untreated (C) or fixed after incubation with DMSO (vehicle) or 20 μM AIM-100 at 37 °C for 2 hours (B). Individual confocal sections from 3D images of YFP fluorescence are shown. Images are representative of at least 3 experiments with each mutant. Scale bars, 10 μm. (D) wt DAT (WT), and YFP-ΔN-DAT mutant expressing cells were incubated with DMSO (vehicle) or 20 μM AIM-100 for 2 hours at 37 °C and lysed. Western blotting of cell lysates was performed using GFP antibody. The results are representative of 2 experiments.