Figure 9.

RCP knock-down increases lysosomal degradation of TfR but not EGFR. (A and C) HeLa cells were either mock transfected or transfected with RCP or Rip11 siRNAs. After 74-h incubation, cells were either permeabilized with saponin to measure total TfR (A) or nonpermeabilized to measure cell surface TfR (C), stained with anti-TfR-PE antibodies, and quantitated by FACS analysis or Western blotting (A, inset). (B and D) Quantitation of data from A and C. The presented data are means ± SD of three independent experiments. (E and F) Cells were transfected with RCP siRNA, fixed, and stained with anti-RCP (E) and anti-TfR (F) antibodies. Arrow points to a HeLa cell with down-regulated RCP. Bars, 2 μM. (G and H) HeLa cells were either mock transfected or transfected with RCP siRNAs. After 74-h incubation, cells were either permeabilized to measure total EGFR (G) or nonpermeabilized to measure cell surface EGFR (H). The levels of EGFR were quantitated by FACS analysis.