CCL5 phosphorylates the 4E-BP1 repressor of mRNA translation through PI-3′ kinase and mTOR. (A) Activated PB T cells were either left untreated or treated with 10 nM CCL5 for the indicated times. Cells were harvested and lysates resolved by SDS-PAGE and immunoblotted with anti–phospho-4E-BP1 (Thr 37/46) antibody or anti–phospho-4E-BP1 (Thr 70) antibody. Membranes were stripped and reprobed for 4E-BP1 as a loading control. The relative fold increase of 4E-BP1 phosphorylation is shown as signal intensity over loading control to the right of each blot. (B) Activated PB T cells were pretreated with either DMSO (carrier), 20 μM LY294002, or 50 nM rapamycin for 1 hour prior to 15-minute treatment with 10 nM CCL5. Cells were harvested and lysates resolved by SDS-PAGE and immunoblotted with anti–phospho-4E-BP1 (Thr 37/46) antibody. Membranes were stripped and reprobed for 4E-BP1 as a loading control. The relative fold increase of 4E-BP1 phosphorylation is shown as signal intensity over loading control. Data are representative of 2 independent experiments plus or minus SD.