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Fig. 3. Ubiquitin ligase activity of c-Cbl determines EGF-induced recruitment of Eps15 towards plasma membrane and endosomes. CHO cells co-expressing FLAG-Eps15, wild-type EGFR and HA-c-Cbl (A-L) or HA-70Z-Cbl (M-X) were incubated with EGF-TxR (red) (A,E,I,M,Q,U) on ice for 1 hour. After stimulation at 37°C for 0 minutes (A-D,M-P), 1 minute (E-H,Q-T) or 5 minutes (I-L,U-X), cells were fixed and stained with anti-FLAG mAb and FITC-conjugated secondary antibody to detect Eps15 proteins (green) (B,F,J,N,R,V) and with anti-HA mAb and Cy5-conjugated secondary antibody to detect Cbl proteins (blue) (C,G,K,O,S,W). Cells were analyzed by confocal microscopy. The displayed confocal planes are from the basal half of the cells.





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