|
|
|
|
|
|
|
|
|||||
| Home Help Feedback Subscriptions Archive Search Table of Contents | |||||
|
Fig. 9. Activation of ERK1/2 and p38 MAP kinases. (A) Top panel, K562 cells were serum-starved, treated with anti-TfR2 mAb G/14C2, with human transferrin (h-Tf) or with anti-TfR1 mAb for the indicated times and subjected to immunoblotting with anti-phospho-ERK1/2 Ab (pERK1/2). Middle panel: in control experiments, serum-starved K562 cells were exposed to anti-TfR1 Ab (5 minutes, 37°C), bovine serum albumin (BSA; 30 minutes, 37°C), bovine transferrin (b-Tf; 30 µM, 30 minutes, 37°C) and phorbol 12-myristate 13-acetate (PMA; 5 minutes, 37°C). Bottom panel, dose-response curve of ERK1/2 phosphorylation over a physiological and sub-physiological range of h-Tf and b-Tf concentrations. Blots were stripped and reprobed for ERK1/2 to ensure equivalent loading and transfer. (B) A TfR2-negative subclone of K562 cell line was serum-starved, treated with h-Tf (30 µM; 30 minutes, 37°C) and PMA (5 minutes, 37°C) and immunoblotted for TfR1, TfR2, pERK1/2 and actin. (C) Top panel, comparison of the level of ERK1/2 phosphorylation in K562 wt and caveolin-1 transfected K562 cells (K562 cav) exposed to anti-TfR2 (5 minutes, 37°C), h-Tf (30 µM; 30 minutes, 37°C) and PMA (5 minutes, 37°C). Bottom panel, serum-starved K562 cells were treated with anti-TfR2 or h-Tf and used for western analysis to assess the phosphorylation of p38 MAP kinase; blots were stripped and reprobed for tubulin to ensure equivalent loading and transfer. For each treatment, the level of ERK1/2 phosphorylation was quantified by densitometry with the Quantity One program (Bio-Rad) and reported in arbitrary units. Error bars show the range of values obtained in three independent experiments.
|
