Fig. 8. BFA induces transport of Tf to apical endosomes associated with Rab25 and Rab11a. (A,B) Image volumes of living cells stably expressing GFP-Rab25 incubated to steady state with basolateral Alexa568-IgA were collected, and reproduced as extended focus projections. IgA and Rab25 closely colocalize, in particular Rab25 is strongly associated with the ARE in which IgA accumulates (arrowheads). (C,D) Image volumes collected of living cells incubated with Alexa568-Tf show that Tf is absent from AREs, so that the apical GFP-Rab25 concentrations correlate with holes in the projected Tf images (arrowheads). (E,F) Images of an apical plane of cells pretreated with BFA for 20 minutes, then incubated with Alexa568-Tf in the continued presence of BFA for another 30 minutes show that BFA induces redirection of Tf into the apical compartments associated with GFP-Rab25. (G,H) Extended focus images of living cells expressing GFP-Rab11a show that, similar to Rab25, Rab11a closely codistributes with basolaterally internalized Alexa568-IgA, particularly in AREs, which again appear as holes in extended focus images of Tf (I,J). (K,L) An apical plane of GFP-Rab11a-expressing cells pretreated with BFA for 20 minutes, then incubated with Alexa568-Tf in the continued presence of BFA for another 30 minutes shows that BFA induces redirection of Tf to the apical compartments associated with GFP-Rab11a. Scale bar, 15 µm (A,B); 10 µm for the remaining panels.
