FIGURE 9.

AdaGb₃ insertion and VT trafficking do not perturb the intracellular traffic of cholera toxin. AdaGb₃ was inserted into HEK-293 cells, and the simultaneous binding and internalization of Alexa488-VTB and Cy3-CTB were assessed. Top panels, binding at 4 °C; bottom panels, detection after 1 h of internalization at 37 °C. DAPI nuclear staining is shown for VT1 B-treated cells without adaGb₃. Bar, 10 μm. Only plasma membrane staining is seen at 4 °C. AdaGb₃-bound VTB and GM1-bound CTB show significant cell surface overlap. At 37 °C, Cy3-CTB is internalized to juxtanuclear Golgi in both control and adaGb₃-treated cells. In contrast, Alexa488-VTB is internalized to punctate vesicles distinct from CTB-labeled Golgi in adaGb₃-treated cells.