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Fig. 4.
Influence of bafilomycin, nocodazole and low temperature on
endocytic pathways in HeLa cells. The transferrin recycling pathway and
the transport of fluid-phase marker to lysosomes is shown. Iron-loaded
transferrin binds to its receptor at the plasma membrane. The complex is
internalized via clathrin-coated vesicles (CCV) and delivered into early endo-
somes within two to five minutes, where the iron is released and transferred
into the cytoplasm. Apotransferrin remains bound to the receptor and recy-
cles via a fast and slow pathway. At the plasma membrane, apotransferrin is
released at the neutral pH. The main route of a fluid-phase marker (e.g.
dextran) from early endosomes (within five min) via endosomal carrier vesi-
cles (ECV) and late endosomes (within 15 min) to lysosomes (within 25
min) is shown in dark grey. Transferrin transport to and recycling via the
PNRC is blocked by nocodazole, whereas bafilomycin and reduced tem-
perature (20
C) are without effect. In contrast, bafilomycin arrests fluid-
phase marker in early endosomes by preventing budding of ECV, whereas
nocodazole leads to accumulation of cargo in ECV. Finally, incubation at
20
°
C prevents delivery of markers from late endosomes to lysosomes. The
endosomal pH values shown are derived from FACS analysis of FITC/
Cy5-dextran and FITC/Cy5-transferrin labeled compartments as well as
from determination of the conformational change of HRV2 occurring at a
threshold of pH
°
≤
5.6.
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