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Fig. 2. Microtubule integrity is required for the ß2-AR trafficking induced by stimulation with isoproterenol: effect of nocodazole. A431 cells stably expressing GFP-tagged ß2AR were stimulated with either 100 nM insulin (+Ins) or 10 µM isoproterenol (+Iso) for 30 minutes (A). Fixed (2% paraformaldehyde in PBS, pH 7.2) cells were immunostained with monoclonal antibodies against {alpha}-tubulin conjugated with Cy3 and examined by confocal microscopy (`microtubules'). The distribution of the GFP-tagged ß2AR and microtubules was analyzed simultaneously by confocal microscopy ((`beta2-AR'). Merging the results from these two analyzes (GFP-receptor in green, microtubules in red; (`merge(') provides a detailed image of the receptor localization. Treatment of the cells with nocodazole (B) induced tubulin depolymerization and blocked the sequestration of the ß2ARs in response to isoproterenol, but not to insulin. Treatment of the cells with latrunculin A (C) blocked sequestration of ß2AR in response to insulin, but not to isoproterenol. Bars, 10 µm.





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