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Fig. 4. Nascent assembly of fluorescent-protein-tagged Cx26 and Cx43 follow the same kinetics and trafficking routes as untagged connexins. BICR-M1R_k cells stably expressing Cx26-YFP or Cx43-GFP were treated with BFA for 6 hours and fixed at different times after BFA removal. In untreated cells, Cx26-YFP and Cx43-GFP exhibited a paranuclear and cell surface localization pattern (A,B). Both Cx26-YFP and Cx43-GFP were localized within the ER-like compartment at 6 hours of BFA treatment and gap junction plaques were absent (C,D). At 30 minutes post-BFA treatment, both Cx26-YFP and Cx43-GFP were localized to the reformed Golgi apparatus (E,F) and by 60 minutes they were also detected at the cell surface as a diffuse rim of fluorescence (G,H). At 90 minutes, both connexins were visualized as cell surface gap junctions while prominent paranuclear populations of both connexins remained (I,J). Two hours after BFA washout, Cx26-YFP and Cx43-GFP localization resembled that of untreated cells (K,L). Bar, 10 µm.

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