FIG. 6.
IL-7 regulates Bim at the posttranslational level. (A) D1 cells were cultured in the present or absence of IL-7. Total-cell extracts were subjected to 2D gel electrophoresis and immunoblotting with anti-Bim. (B) Cell extracts were prepared from D1 cells cultured with or without IL-7, immunoprecipitated with anti-Bim, and treated with CIAP, followed by blotting with anti-phosphoserine 4A4 and anti-Bim. (C) D1 cells were infected with retroviruses expressing either GFP alone (pMIG), WT BimEL, or mutants of BimEL. The infected cells were cultured at 37°C for 16 h in the present of 50 ng/ml of IL-7. Apoptotic cells were determined by annexin V/7-AAD staining. The proportion of apoptotic cells was calculated as the percentage of vector (pMIG)-infected cells (GFP-positive cells) that were positive for annexin V. Values are means ± standard deviations for three experiments (*, P = 0.01; **, P = 0.042; ***, P ≤ 0.0098).