Fig. 1. DynK44A inhibits CTX internalization into endosomes and the Golgi apparatus. FITC-CTX internalized for 30 minutes at 37°C (A,D) is localized to both endosomes labeled for TfR (B) and the Golgi apparatus labeled for GM130 (E). The merged confocal images present FITC-CTX in green (C,F) and TfR (C) or GM130 (F) in red and colocalization in yellow. Alternatively, NIH-3T3 cells were infected with both the tTA and dynK44A adenoviruses (G-J), and after 36 hours, the cells were coincubated with FITC-CTX (H) and Rh-Tf (I) for 30 minutes at 37°C prior to fixation. HA-tagged dynK44A-infected cells (arrows) were identified by postfixation labeling with anti-HA antibodies followed by Alexa647 anti-mouse secondary antibodies (G). The merged confocal image presents FITC-CTX in green, Rh-Tf in red and colocalization in yellow (J). Dramatic reduction in CTX and Rh-Tf internalization was observed in the dynK44A expressing cell. Bar, 20 µm.

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