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Fig. 8. Upregulation of TfR expression. Bloodstream 221 cells were pulse-chase radiolabeled as in Fig. 7 in the absence (A) or presence (B) of FMK024, and cell and media fractions were prepared at the indicated times. TfR polypeptides were immunoprecipitated with anti-TfR and analyzed by SDS-PAGE and phosphorimaging. (C) Bloodstream 221 cells were cultured 48 hours in HMI9 media containing 10% fetal bovine serum (FBS), 10% canine serum (CS), or 10% canine serum plus 200 µg/ml bovine transferrin (CS⁺). Cells were then pulse labeled (15 minutes) and chased under the same conditions. Cell (c) and media (m) fractions were prepared at 0 and 4 hours and analyzed as above. Fractional recovery of released ESAG7 polypeptide for each set is presented below (boxed). The relative amounts of initial ESAG7 are 1.0:14.0:6.1 for FBS (lane 1):CS (lane 5):Cs⁺ (lane 9). Distortions of lanes 5-12 are reproducible and probably result from non-specific precipitation of components of dog serum in lanes 6, 8, 10 and 12. (A-C) Mobilities of mature and immature forms of ESAG6 (E6m and E6i) and ESAG7 (E7) are indicated on the left. Mobilities of molecular mass standards are indicated on the right. All lanes contain 10⁷ cell equivalents.

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