Stimulation of CD74 by anti-CD74 or MIF in normal and malignant cells induces BCL-X_L expression. (A-B) BCL-X_L expression. IgD⁺ B cells from control mice or B cells from CD74^–/– mice were stimulated with anti-CD74 antibody or nonspecific anti-CD8 antibody for 24 hours. Total RNA was isolated, and reverse transcription was carried out using Superscript II RT. The results presented are representative of 5 separate experiments (A). 293 cells transfected with the FL CD74 construct. Five hours following transfection the cells were stimulated or not with anti-CD74 or nonspecific antibodies (anti-CD8) for 12 hours (B). RT-PCR was performed as described in “Materials and methods.” (C) 293 cells transfected with the CD74 1-82 or 1-82 mutated in its 42 to 44 aa (1-82*42-44) constructs. Five hours following transfection the cells were stimulated or not with anti-CD74 for 12 hours. Reverse transcriptase–polymerase chain reaction (RT-PCR) was performed as described in “Materials and methods.” (D) B cells from control or CD74^–/– mice were stimulated in the presence or absence of MIF for 10 hours as described in “Materials and methods.” Total RNA was isolated, and reverse transcription was carried out using Superscript II RT. The results presented are representative of 3 separate experiments. (E) Stage I and stage IV B-CLL B cells were stimulated in the presence or absence of anti-CD74 antibody or nonspecific anti-ID2 antibody for 18 hours. Total RNA was isolated, and reverse transcription was carried out using Superscript II RT. The results presented are representative of 6 separate experiments. The intensity of the BCL-X_L band following each treatment was divided by the intensity of the HPRT band in each treatment. The activation-fold ratio in the absence of any treatment was normalized to 1, and the ratio for each treatment was calculated as the intensity of the treatment sample relative to 1.