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. Author manuscript; available in PMC: 2011 Mar 1.
Published in final edited form as: Cytoskeleton (Hoboken). 2010 Mar;67(3):193–206. doi: 10.1002/cm.20437

Figure 8.

Figure 8

WASH depletion or disruption of the actin cytoskeleton delays EGF transport to LAMP1-positive late endosomes. (A) Fibroblast-like cells (FLCs) were transfected with nonspecific siRNA (control) or WASH siRNA, or treated with latrunculin A (Lat A) or cytochalasin D (Cyto D). After Alexa Fluor 488 EGF complex (green) was internalized for 10 min at 37 °C and chased for 30 min at 37 °C, cells were fixed, and LAMP1 (red) was stained by immunofluorescence and DNA (blue) was stained with DAPI. Scale bar, 5 μm. (B) Extracts from FLCs transfected with control or WASH siRNAs were separated by SDS-PAGE and immunoblotted with anti-WASH or anti-GAPDH antibodies. (C) The amount of fluorescent EGF colocalized with LAMP1-positive vesicles as a percentage of the total fluorescent EGF is shown for each treatment. 10 cells were counted for each condition, and data are mean ± SD of three independent experiments. Asterisk (*) indicates a p-value < 0.05.