| |
Volume 1 Issue 2 February 2000
Default membrane-volume sorting is followed by polar sorting in compartments preceding the apical recycling endosome of MDCK cells
Paul S. Brown, Exing Wang, Benjamin Aroeti, Steven J. Chapin, Keith E. Mostov and Kenneth W. Dunn
Figure 2 - Tf, LDL and IgA are internalized into the same lateral sorting endosomes. Filter grown PTR9 cells were incubated from the basolateral side for 2 minutes at 37°C with both diI-LDL and F-Tf (panels A - B, respectively) or with both TxR-Tf and OG-IgA (panels C - D, respectively) and images collected by confocal microscopy. Although the ratio of ligands varies between endosomes, extensive colocalization is seen between each pair of probes. Panel E shows a stereopair projection of a field of cells fixed after a 2 minute basolateral incubation with F-Tf. In this and all following stereopair images, the apical side of the cells will appear towards the viewer when using a stereo-viewer, but will appear away from the viewer when viewed cross-eyed. Note that this and all following stereopairs are represented as movies of rotated projections, click on the icons below or visit the website located at http://renal.nephrology.iupui.edu/brownetal. Panels F and G show fields of cells incubated for 60 minutes at 4°C with diI-LDL and F-Tf, respectively. Length of the scale bar reflects 5 microns in panels A-D and 7 microns in panels E-G.
Figure 7 - IgA initially colocalizes with Tf, but is subsequently sorted to an apical compartment depleted in Tf. A. Stereopair images of cells fixed after incubation with TxR-Tf (red) and OG-IgA (green) for 10 minutes show an extensive and proportional colocalization of Tf and IgA throughout the cells. B. Cells incubated for 15 minutes with OG-IgA (green) and TxR-Tf (red) demonstrate that IgA is subsequently sorted from Tf to an apical compartment. C. Cells labeled for 15 minutes with OG-Tf (green) and TxR-IgA (red) after prior treatment for 60 minutes in 33 mM nocodazole show no apical and axial transport of either Tf or IgA. D. Cells incubated with OG-Tf (green) for 15 minutes, with TxR-IgA (red) included for the last 5 minutes show no indication of an apical, IgA-enriched compartment. E. Cells incubated for 15 minutes with OG-Tf (green), with TxR-IgA (red) included for 5 minutes, then chased for 5 minutes show that during the 5 minute chase IgA is sorted to the apical compartment. Scale bars were omitted from this figure as they interfere with stereo image formation. The field in panel A is 47 microns in height, B and C are 38 mM in height, D and E are 25 mM in height.
Note that these stereopair figures are represented as movies of rotated projections. You can click the icons below or visit the website located at http://renal.nephrology.iupui.edu/brownetal
|
|
